Use of 1H-indazole-3-carboxamide compounds as glycogen synthase kinase 3 beta inhibitors

ABSTRACT

1H-indazole-3-carboxamide compounds as glycogen synthase kinase 3 beta inhibitors. The present invention relates to the 1H-indazole-3-carboxamide compounds for use as glycogen synthase kinase 3 beta (GSK-3β) inhibitors and to their use in the treatment of GSK-3β-related disorders such as, for example, (i) insulin-resistance disorders; (ii) neurodegenerative diseases; (iii) mood disorders; (iv) schizophrenic disorders; (v) cancerous disorders; (vi) inflammation, (vii) substance abuse disorders; and (viii) epilepsies.

FIELD OF THE INVENTION

The present invention relates to the new use of1H-indazole-3-carboxamide compounds acting as glycogen synthase kinase 3beta (GSK-3β) inhibitors and to their use in the treatment ofGSK-3β-related disorders such as (i) insulin-resistance disorders; (ii)neurodegenerative diseases; (iii) mood disorders; (iv) schizophrenicdisorders; (v) cancerous disorders; (vi) inflammation, (vii) substanceabuse disorders; and (viii) epilepsies.

STATE OF THE ART

Protein kinases constitute a large family of structurally relatedenzymes, which transfer phosphate groups from high-energy donormolecules (such as adenosine triphosphate, ATP) to specific substrates,usually proteins. After phosphorylation, the substrate undergoes to afunctional change, by which kinases can modulate various biologicalfunctions.

In general, protein kinases can be divided in several groups, accordingto the substrate that is phosphorylated. For example, serine/threoninekinase phosphorylates the hydroxyl group on the side chain of serine orthreonine aminoacid.

Glycogen synthase kinases 3 (GSK-3) are constitutively activemultifunctional enzymes, quite recently discovered, belonging to theserine/threonine kinases group.

Human GSK-3 are encoded by two different and independent genes, whichleads to GSK-3α and GSK-3β proteins, with molecular weights of about 51and 47 kDa, respectively. The two isoforms share nearly identicalsequences in their kinase domains, while outside of the kinase domain,their sequences differ substantially (Benedetti et al., NeuroscienceLetters, 2004, 368, 123-126). GSK-3α is a multifunctional protein serinekinase and GSK-3β is a serine-threonine kinase.

It has been found that GSK-3β is widely expressed in all tissues, withwidespread expression in the adult brain, suggesting a fundamental rolein neuronal signaling pathways (Grimes and Jope, Progress inNeurobiology, 2001, 65, 391-426). Interest in glycogen synthase kinases3 arises from its role in various physiological pathways, such as, forexample, metabolism, cell cycle, gene expression, embryonic developmentoncogenesis and neuroprotection (Geetha et al, British JournalPharmacology, 2009, 156, 885-898).

GSK-3β was originally identified for its role in the regulation ofglycogen synthase for the conversion of glucose to glycogen (Embi etal., Eur J Biochem, 1980, 107, 519-527). GSK-3β showed a high degree ofspecificity for glycogen synthase.

Type 2 diabetes was the first disease condition implicated with GSK-3β,due to its negative regulation of several aspects of insulin signalingpathway. In this pathway 3-phosphoinositide-dependent protein kinase 1(PDK-1) activates PKB, which in turn inactivates GSK-3β. Thisinactivation of GSK-3β leads to the dephosphorylation and activation ofglycogen synthase, which helps glycogen synthesis (Cohen et al., FEBSLett., 1997, 410, 3-10). Moreover, selective inhibitors of GSK-3β areexpected to enhances insulin signaling in prediabetic insulin-resistantrat skeletal muscle, thus making GSK-3β an attractive target for thetreatment of skeletal muscle insulin resistance in the pre-diabeticstate (Dokken et al., Am J. Physiol. Endocrinol. Metab., 2005, 288,E1188-E1194).

GSK-3β was also found to be a potential drug target in otherspathological conditions due to insulin-resistance disorders, such assyndrome X, obesity and polycystic ovary syndrome (Ring D B et al.,Diabetes, 2003, 52: 588-595).

It has been found that GSK-3β is involved in the abnormalphosphorylation of pathological tau in Alzheimer's disease (Hanger etal., Neurosci. Lett., 1992, 147, 58-62; Mazanetz and Fischer, Nat RevDrug Discov., 2007, 6, 464-479; Hong and Lee, J. Biol. Chem., 1997, 272,19547-19553). Moreover, it was proved that early activation of GSK-3β,induced by apolipoprotein ApoE4 and β-amyloid, could lead to apoptosisand tau hyperphosphorylation (Cedazo-Minguez et al., Journal ofNeurochemistry, 2003, 87, 1152-1164). Among other aspect of Alzheimer'sdisease, it was also reported the relevance of activation of GSK-3β atmolecular level (Hernandez and Avila, FEBS Letters, 2008, 582,3848-3854).

Moreover, it was demonstrated that GSK-3β is involved in the genesis andmaintenance of neurodegenerative changes associated with Parkinson'sdisease (Duka T. et al., The FASEB Journal, 2009; 23, 2820-2830).

Accordingly to these experimental observations, inhibitors of GSK-3β mayfind applications in the treatment of the neuropathological consequencesand the cognitive and attention deficits associated with tauopathies;Alzheimer's disease; Parkinson's disease; Huntington's disease (theinvolvement of GSK-3β in such deficits and diseases is disclosed inMeijer L. et al., TRENDS Pharm Sci, 2004; 25, 471-480); dementia, suchas, but not limited to, vascular dementia, post-traumatic dementia,dementia caused by meningitis and the like; acute stroke; traumaticinjuries; cerebrovascular accidents; brain and spinal cord trauma;peripheral neuropathies; retinopathies and glaucoma (the involvement ofGSK-3β in such conditions is disclosed in WO 2010/109005).

The treatment of spinal neurodegenerative disorders, like amyotrophiclateral sclerosis, multiple sclerosis, spinal muscular atrophy andneurodegeneration due to spinal cord injury has been also suggested inseveral studies related to GSK-3β inhibition, such as, for example inCalderó J. et al., “Lithium prevents excitotoxic cell death ofmotoneurons in organotypic slice cultures of spinal cord”, Neuroscience.2010 Feb. 17; 165(4):1353-69, Léger B. et al., “Atrogin-1, MuRF1, andFoXO, as well as phosphorylated GSK-3beta and 4E-BP1 are reduced inskeletal muscle of chronic spinal cord-injured patients”, Muscle Nerve,2009 July; 40(1):69-78, and Galimberti D. et al., “GSK3β geneticvariability in patients with Multiple Sclerosis”, Neurosci Lett. 2011Jun. 15; 497(1):46-8.

Furthermore, GSK-3β has been linked to the mood disorders, such asbipolar disorders, depression, and schizophrenia.

Inhibition of GSK-3β may be an important therapeutic target of moodstabilizers, and regulation of GSK-3β may be involved in the therapeuticeffects of other drugs used in psychiatry. Dysregulated GSK-3β in mooddisorder, bipolar disorder, depression and schizophrenia could havemultiple effects that could impair neural plasticity, such as modulationof neuronal architecture, neurogenesis, gene expression and the abilityof neurons to respond to stressful, potentially lethal conditions (Jopeand Roh, Curr. Drug Targets, 2006, 7, 1421-1434).

The role of GSK-3β in mood disorder was highlighted by the study oflithium and valproate (Chen et al., J. Neurochem., 1999, 72, 1327-1330;Klein and Melton, Proc. Natl. Acad. Sci. USA, 1996, 93, 8455-8459), bothof which are GSK-3β inhibitors and are used to treat mood disorders.There are also existing reports from the genetic perspective supportingthe role of GSK-3β in the disease physiology of bipolar disorder (Gould,Expert. Opin. Ther. Targets, 2006, 10, 377-392).

It was reported a decrease in AKT1 protein levels and itsphosphorylation of GSK-3β at Serine-9 in the peripheral lymphocytes andbrains of individuals with schizophrenia. Accordingly, this findingsupports the proposal that alterations in AKT1-GSK-3β signalingcontribute to schizophrenia pathogenesis (Emamian et al., Nat Genet,2004, 36, 131-137).

Additionally, the role of GSK-3β in cancer is a well-acceptedphenomenon.

The potential of small molecules that inhibit GSK-3β has been evidencedfor some specific cancer treatments (Jia Luo, Cancer Letters, 2009, 273,194-200). GSK-3β expression and activation are associated with prostatecancer progression (Rinnab et al., Neoplasia, 2008, 10, 624-633) and theinhibition of GSK3b was also proposed as specific target for pancreaticcancer (Garcea et al., Current Cancer Drug Targets, 2007, 7, 209-215)and ovarian cancer (Qi Cao et al., Cell Research, 2006, 16 671-677).Acute inhibition of GSK-3β in colon-rectal cancer cells activatesp53-dependent apoptosis and antagonizes tumor growth (Ghosh et al., ClinCancer Res 2005, 11, 4580-4588).

The identification of a functional role for GSK-3β in MLL-associatedleukaemia suggests that GSK-3β inhibition may be a promising therapythat is selective for transformed cells that are dependent on HOXoverexpression (Birch et al., Cancer Cell, 2010, 17, 529-531).

GSK-3β is involved in numerous inflammatory signalling pathways, forexample, among others GSK-3β inhibition has been shown to inducesecretion of the anti-inflammatory cytokine IL-10. According to thisfinding, GSK-3β inhibitors could be useful to regulate suppression ofinflammation (G. Klamer et al., Current Medicinal Chemistry, 2010,17(26), 2873-2281, Wang et al., Cytokine, 2010, 53, 130-140).

GSK-3β inhibition has been also shown to attenuate cocaine-inducedbehaviors in mice. The administration of cocaine in mice pretreated witha GSK-3β inhibitor demonstrated that pharmacological inhibition of GSK3reduced both the acute behavioral responses to cocaine and the long-termneuroadaptations produced by repeated cocaine (Cocaine-inducedhyperactivity and sensitization are dependent on GSK3, Miller J S et al.Neuropharmacology. 2009 June; 56(8):1116-23, Epub 2009 Mar. 27).

The role of GSK-3β in the development of several forms of epilepsies hasbeen demonstrated in several studies, which suggest that inhibition ofGSK-3β could be a pathway for the treatment of epilepsy (Novel glycogensynthase kinase 3 and ubiquitination pathways in progressive myoclonusepilepsy, Lohi H et al., Hum Mol Genet. 2005 Sep. 15; 14(18):2727-36 andHyperphosphorylation and aggregation of Tau in laforin-deficient mice,an animal model for Lafora disease, Puri R et al., J Biol Chem. 2009Aug. 21; 284(34):22657-63).

The relationship between GSK-3β inhibition and treatment of pain hasbeen demonstrated by Martins D F et al. in “The antinociceptive effectsof AR-A014418, a selective inhibitor of glycogen synthase kinase-3 beta,in mice”, J. Pain, 2011 March; 12(3):315-22.

A review on GSK-3β, its function, its therapeutic potential and itspossible inhibitors is given in “GSK-3β: role in therapeutic landscapeand development of modulators” (S. Phukan et al., British Journal ofPharmacology (2010), 160, 1-19).

WO 2004/014864 discloses 1H-indazole-3-carboxamide compounds asselective cyclin-dependant kinases (CDK) inhibitors. Such compounds areassumed to be useful in the treatment of cancer, through a mechanismmediated by CDK₂, and neurodegenerative diseases, in particularAlzheimer's disease, through a mechanism mediated by CDK₅, and asanti-viral and anti-fungine, through a mechanism mediated by CDK₇, CDK₈and CDK₉.

Cyclin-dependant kinases (CDKs) are serine/threonine kinases, firstdiscovered for their role in regulating the cell cycle. CDKs are alsoinvolved in regulating transcription, mRNA processing, and thedifferentiation of nerve cells. Such kinases activate only after theirinteraction and binding with regulatory subunits, namely cyclins.

Moreover, 1H-indazole-3-carboxamide compounds were also described asanalgesics in the treatment of chronic and neuropathic pain (see, forexample, WO 2004/074275 and WO 2004/101548) and as 5-HT₄ receptorantagonists, useful in the treatment of gastrointestinal disorders,central nervous system disorders and cardiovascular disorders (see, forexample, WO 1994/10174).

SUMMARY OF THE INVENTION

As GSK-3β had been only recently discovered as a pharmacological target,there is a strong need to find compounds that selectively inhibitsGSK-3β.

The Applicant has surprisingly found that the 1H-indazole-3-carboxamidecompounds of formula (I) are capable of inhibit GSK-3β and have veryhigh affinity for GSK-3β, when compared with other kinases. Thus, saidcompounds are capable of selectively inhibiting GSK-3β.

Accordingly, the useful compounds according to this invention arecapable of selectively inhibiting the activity of GSK-3β and are,therefore, useful for the treatment of the pathological conditionsarising from the uncontrolled activation and/or over-expression ofGSK-3β, selected from the group comprising (i) insulin-resistancedisorders, such as type-2 diabetes, syndrome X, obesity and polycysticovary syndrome; (ii) neurodegenerative diseases, such as Parkinson'sdisease, Alzheimer's disease, Huntington's disease and spinalneurodegenerative disorders; (iii) mood disorders, such as bipolardisorders and depressive disorders; (iv) schizophrenic disorders; (v)cancerous disorders, such as prostate, pancreatic, ovarian, andcolon-rectal cancer and MLL-associated leukaemia; (vi) inflammation;(vii) substance abuse disorders; and (viii) epilepsies.

Then, in a first aspect, the present invention relates to the use of1H-indazole-3-carboxamide compounds having the following general formula(I)

-   -   wherein    -   R_(a) and R_(a)′, equal or different each other, is a hydrogen        atom; a halogen atom; a C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆        alkynyl and C₁-C₆ alkoxy group, optionally substituted by one or        more substituents selected from the group consisting of halogen,        hydroxy, —NH₂, and C₁-C₃ alkoxy; a carbocyclic or heterocyclic        ring, aliphatic or aromatic, having from 3 to 12 members,        optionally substituted by one or more substituents selected from        the group consisting of halogen, hydroxy, C₁-C₆ alkyl, C₁-C₆        alkoxy, —NR₁R₂, —C(O)OH, —C(O)OR₁ and —C(O)NR₁R₂;    -   Y is a bond, a C₁-C₆ alkyl, C₂-C₆ alkenyl or C₂-C₆ alkynyl        group, optionally substituted by one or more substituents        selected from the group consisting of halogen, hydroxy, —NH₂,        and C₁-C₃ alkoxy;    -   R_(b) is a carbocyclic or heterocyclic ring, aliphatic or        aromatic, having from 3 to 12 members, substituted by one or        more substituents selected from the group consisting of halogen,        hydroxy, nitro, cyano, —CF₃, C₁-C₆ alkoxy, benzyloxy, C₁-C₄        alkyl, C₂-C₄ alkenyl, and C₂-C₄ alkynyl, —NHSO₂CH₃, —SO₂NH₂,        —Z—C(O)OH, —Z—C(O)OR₁ and —Z—C(O)NR₁R₂, wherein Z is a σ-bond or        (C₁-C₃)alkyl;    -   R₁ and R₂ are independently a hydrogen atom, a C₁-C₄ alkyl        group, a C₂-C₄ alkenyl group, a C₂-C₄ alkynyl group, and a        phenyl group;    -   and its salts of addition with pharmaceutically acceptable        organic and inorganic acids and bases;    -   for the treatment of a disease arising from the uncontrolled        activation and/or over-expression of GSK-3β, selected from the        group consisting of (i) insulin-resistance disorders, such as        type-2 diabetes, syndrome X, obesity and polycystic ovary        syndrome; (ii) neurodegenerative diseases, such as Parkinson's        disease, Alzheimer's disease, Huntington's disease and spinal        neurodegenerative disorders; (iii) mood disorders, such as        bipolar disorders and depressive disorders; (iv) schizophrenic        disorders; (v) cancerous disorders, such as prostate,        pancreatic, ovarian, and colon-rectal cancer and MLL-associated        leukaemia; (vi) inflammation; (vii) substance abuse disorders;        and (viii) epilepsies.

In a second aspect, the present invention relates to a method oftreatment of a pathological state arising from the uncontrolledactivation and/or over-expression of GSK-3β, selected from the groupconsisting of (i) insulin-resistance disorders, such as type-2 diabetes,syndrome X, obesity and polycystic ovary syndrome; (ii)neurodegenerative diseases, such as Parkinson's disease, Alzheimer'sdisease, Huntington's disease and spinal neurodegenerative disorders;(iii) mood disorders, such as bipolar disorders and depressivedisorders; (iv) schizophrenic disorders; (v) cancerous disorders, suchas prostate, pancreatic, ovarian, and colon-rectal cancer andMLL-associated leukaemia; (vi) inflammation; (vii) substance abusedisorders; and (viii) epilepsies by the administration to a human beingin need thereof of an effective amount of a 1H-indazole-3-carboxamidehaving the following general formula (I)

-   -   wherein    -   R_(a) and R_(a)′, equal or different each other, is a hydrogen        atom; a halogen atom; a C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆        alkynyl and C₁-C₆ alkoxy group, optionally substituted by one or        more substituents selected from the group consisting of halogen,        hydroxy, —NH₂, and C₁-C₃ alkoxy; a carbocyclic or heterocyclic        ring, aliphatic or aromatic, having from 3 to 12 members,        optionally substituted by one or more substituents selected from        the group consisting of halogen, hydroxy, C₁-C₆ alkyl, C₁-C₆        alkoxy, —NR₁R₂, —C(O)OH, —C(O)OR₁ and —C(O)NR₁R₂;    -   Y is a bond, a C₁-C₆ alkyl, C₂-C₆ alkenyl or C₂-C₆ alkynyl        group, optionally substituted by one or more substituents        selected from the group consisting of halogen, hydroxy, —NH₂,        and C₁-C₃ alkoxy;    -   R_(b) is a carbocyclic or heterocyclic ring, aliphatic or        aromatic, having from 3 to 12 members, substituted by one or        more substituents selected from the group consisting of halogen,        hydroxy, nitro, cyano, —CF₃, C₁-C₆ alkoxy, benzyloxy, C₁-C₄        alkyl, C₂-C₄ alkenyl, and C₂-C₄ alkynyl, —NHSO₂CH₃, —SO₂NH₂,        —Z—C(O)OH, —Z—C(O)OR₁ and —Z—C(O)NR₁R₂, wherein Z is a σ-bond or        (C₁-C₃)alkyl;    -   R₁ and R₂ are independently a hydrogen atom, a C₁-C₄ alkyl        group, a C₂-C₄ alkenyl group, a C₂-C₄ alkynyl group, and a        phenyl group;    -   and its salts of addition with pharmaceutically acceptable        organic and inorganic acids and bases.

The present invention also includes the prodrugs, stereoisomers, andenantiomers of the compounds of formula (I) described above.

Some compounds falling within the above formula (I) are new, i.e., werenever disclosed and exemplified in a printed publication before the dateof filing of the present application.

Accordingly, in a third aspect, the present invention relates to1H-indazole-3-carboxamides compounds of formula:

-   N-{[1-(2,4-dichlorobenzyl)piperidin-4-yl]methyl}-5-methoxy-1H-indazole-3-carboxamide    and-   N-({1-[4-(benzyloxy)benzyl]piperidin-4-yl}methyl)-5-methoxy-1H-indazole-3-carboxamide.

DETAILED DESCRIPTION OF THE INVENTION

Throughout the present description and the following claims, “C₁₋₆alkyl” is intended to indicate linear or branched alkyl groups havingfrom 1 to 6 carbon atoms, such as methyl, ethyl, propyl, isopropyl,n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, sec-pentyl,3-pentyl, iso-pentyl, neo-pentyl, n-hexyl, sec-hexyl and neo-hexyl.

Throughout the present description and the following claims, “C₁₋₄alkyl” is intended to indicate linear or branched alkyl groups havingfrom 1 to 4 carbon atoms, such as methyl, ethyl, propyl, isopropyl,n-butyl, isobutyl, sec-butyl and tert-butyl.

Throughout the present description and the following claims, “C₁₋₃alkyl” is intended to indicate linear or branched alkyl groups havingfrom 1 to 3 carbon atoms, such as methyl, ethyl, propyl and isopropyl.

Throughout the present description and the following claims, “C₂₋₆alkenyl” is intended to indicate linear or branched alkyl groups havingfrom 2 to 6 carbon atoms and at least one double bond, such as ethenyl(vinyl), 1-propenyl, 2-propenyl (allyl), isopropenyl, butenyl, pentenyland hexenyl.

Throughout the present description and the following claims, “C₂₋₄alkenyl” is intended to indicate linear or branched alkyl groups havingfrom 2 to 4 carbon atoms and at least one double bond, such as ethenyl(vinyl), 1-propenyl, 2-propenyl (allyl), isopropenyl and butenyl.

Throughout the present description and the following claims, “C₂₋₆alkynyl” is intended to indicate linear or branched alkyl groups havingfrom 2 to 6 carbon atoms and at least one triple bond, such as ethynyl,1-propynyl, 2-propynyl (propargyl), butynyl, pentynyl and hexynyl.

Throughout the present description and the following claims, “C₂₋₄alkynyl” is intended to indicate linear or branched alkyl groups havingfrom 2 to 4 carbon atoms and at least one triple bond, such as ethynyl,1-propynyl, 2-propynyl (propargyl) and butynyl.

Throughout the present description and the following claims, “C₁₋₆alkoxy” is intended to indicate linear or branched alkoxy groups havingfrom 1 to 6 carbon atoms, such as methoxy, ethoxy, n-propoxy,iso-propoxy, n-butoxy, tert-butoxy, n-penthoxy, sec-penthoxy,isopenthoxy and n-esiloxy.

Throughout the present description and the following claims, “C₁₋₃alkoxy” is intended to indicate linear or branched alkoxy groups havingfrom 1 to 3 carbon atoms, such as methoxy, ethoxy, n-propoxy andiso-propoxy.

According to a preferred embodiment of the invention, the meanings ofR_(a), R_(a)′, R_(b) and Y of the formula (I) above are described herein below.

Preferably, R_(a) and R_(a)′, equal or different each other, is ahydrogen atom; a halogen atom, selected from chlorine, bromine andiodine; a C₁-C₆ alkyl, and C₁-C₆ alkoxy group, optionally substituted byone or more substituents selected from the group consisting of halogen,hydroxy, —NH₂, or C₁-C₃ alkoxy; a carbocyclic or heterocyclic ring,aliphatic or aromatic, having from 4 to 10 members, optionallysubstituted by one or more substituents selected from the groupconsisting of halogen, hydroxy, C₁-C₆ alkyl, C₁-C₆ alkoxy, —NR₁R₂,—C(O)OH, —C(O)OR₁ and —C(O)NR₁R₂.

More preferably, R_(a) and R_(a)′, equal or different each other, is ahalogen atom, selected from chlorine and bromine; a C₁-C₆ alkyl group; aC₁-C₆ alkoxy group; or a carbocyclic or heterocyclic ring, aliphatic oraromatic, having from 5 to 6 members, optionally substituted by one ormore substituents, selected from the group consisting of halogen,hydroxy, C₁-C₆ alkyl, C₁-C₆ alkoxy, —NR₁R₂ and —C(O)OH.

Advantageously, said carbocyclic or heterocyclic ring, aliphatic oraromatic, having 5 or 6 members is selected from phenyl, pyridine,pyrimidine, pyrazine, pyridazine, pyrrole, furan, thiophene, oxazole,isoxazole, thiazole, isothiazole, 2H-pyran, cyclohexyl, cyclopenthylpiperidine, piperazine.

Even more preferably, R_(a) and R_(a)′, equal or different each other,is a bromine atom, a C₁-C₃ alkoxy group; or an aromatic carbocyclic orheterocyclic ring, having 6 members, optionally substituted by one ortwo substituents selected from the group consisting of halogen, hydroxy,C₁-C₃ alkyl, C₁-C₃ alkoxy, —NR₁R₂ and —C(O)OH.

In a preferred embodiment, said carbocyclic or heterocyclic ring,aliphatic or aromatic, having 6 members is selected from phenyl,pyridine, pyrimidine, pyrazine, pyridazine, 2H-pyran, cyclohexyl,piperidine, piperazine.

In an even more preferred embodiment, said carbocyclic or heterocyclicring, aliphatic or aromatic, having 6 members is selected from phenyl,pyridine, pyrimidine, 2H-pyran, cyclohexyl.

Preferably, Y is a bond, C₁-C₆ alkyl group, optionally substituted byone or more substituents selected from the group consisting of halogen,hydroxy, —NH₂, and C₁-C₃ alkoxy.

More preferably, Y is a C₁-C₆ alkyl group.

Even more preferably, Y is a C₁-C₃ alkyl group.

Preferably, R_(b) is a carbocyclic or heterocyclic ring, aliphatic oraromatic, having from 4 to 10 members, substituted by one or moresubstituents selected from the group consisting of halogen, hydroxy,nitro, cyano, —CF₃, C₁-C₆ alkoxy, benzyloxy, C₁-C₄ alkyl, —NHSO₂CH₃,—SO₂NH₂, —Z—C(O)OH, —Z—C(O)OR₁ and —Z—C(O)NR₁R₂, wherein Z is a σ-bondor (C₁-C₃)alkyl.

More preferably, R_(b) is a carbocyclic or heterocyclic ring, aliphaticor aromatic, having from 5 to 6 members, substituted by one or moresubstituents, selected from the group consisting of halogen, hydroxy,nitro, —CF₃, C₁-C₆ alkoxy, benzyloxy, —NHSO₂CH₃, —SO₂NH₂, —Z—C(O)OH and—Z—C(O)OR₁, wherein Z is a σ-bond or (C₁-C₃)alkyl.

Advantageously, said carbocyclic or heterocyclic ring, aliphatic oraromatic, having 5 or 6 members is selected from phenyl, pyridine,pyrimidine, pyrazine, pyridazine, morpholine, pyrrole, furan, thiophene,oxazole, isoxazole, thiazole, isothiazole, 1-oxa-2,4-diazole, 2H-pyran,cyclohexyl, cyclopenthyl piperidine, piperazine.

Even more preferably, R_(b) is an aromatic carbocyclic ring having 6members substituted by one or two substituents selected from the groupconsisting of halogen, hydroxy, nitro, —CF₃, C₁-C₃ alkoxy and benzyloxy.

In a preferred embodiment, said carbocyclic or heterocyclic ring,aliphatic or aromatic, having 6 members is selected from phenyl,pyridine, pyrimidine, pyrazine, pyridazine, morpholine, 2H-pyran,cyclohexyl, piperidine, piperazine.

In an even more preferred embodiment, said carbocyclic or heterocyclicring, aliphatic or aromatic, having 6 members is selected from phenyl,pyridine, pyrimidine, morpholine, 2H-pyran, cyclohexyl.

In an even more preferred embodiment, said carbocyclic or heterocyclicring, aliphatic or aromatic, having 5 members is selected from furan,thiophene, thiazole, oxazole, and 1-oxa-2,4-diazole.

Preferably, R₁ and R₂ are independently a hydrogen atom, a C₁-C₄ alkylgroup, or a phenyl group.

More preferably, R₁ and R₂ are independently a C₁-C₃ alkyl group.

Even more preferably, R₁ and R₂ are both a methyl group.

Preferably, said carbocyclic or etherocyclic ring, aliphatic oraromatic, having from 5 to 6 members is selected from the groupconsisting of phenyl, ciclohexane, ciclopentane, pyridine, pyrazine,pyrimidine, pyridazine, piperidine, piperazine, furan, thiophene,pyrrole, pyrrolidine, imidazole, morpholine, thiazole, thiazolidine,thiadiazole, thiadiazolidine, oxazole, oxazolidine, isoxazole,isoxazolidine, pyrazole.

More preferably, said carbocyclic ring is phenyl and said etherocyclicring is pyridine, oxazole, imidazole and pyrrole.

The compounds useful in the present invention are preferably employed assalts with pharmaceutically acceptable organic and inorganic acids orbases.

Preferably, the pharmaceutically acceptable organic acids are selectedfrom the group consisting of oxalic, maleic, methanesulphonic,paratoluenesulphonic, succinic, citric, malic, tartaric lactic acid.

Preferably, the pharmaceutically acceptable organic bases are selectedfrom the group consisting of tromethamine, lysine, arginine, glycine,alanine and ethanolamine.

Preferably, the pharmaceutically acceptable inorganic acids are selectedfrom the group consisting of hydrochloric, hydrobromic, phosphoric andsulfuric acid.

Preferably, the pharmaceutically acceptable inorganic bases are selectedfrom the group consisting of hydroxide or carbonate of alkaline oralkaline-earth metals, such as sodium, potassium and calcium.

The present invention also includes the use of prodrugs, stereoisomers,and enantiomers of the compounds of formula (I) described above.

As used herein the term “prodrug” refers to an agent, which is convertedinto the parent drug in vivo by some physiological chemical process(e.g., a prodrug on being brought to the physiological pH is convertedto the desired drug form). Prodrugs are often useful because, in somesituations, they may be easier to administer than the parent drug. Theymay, for instance, be bioavailable by oral administration whereas theparent drug is not. The prodrug may also have improved solubility inpharmacological compositions over the parent drug. An example, withoutlimitation, of a prodrug would be a compound of the present inventionwherein it is administered as an ester (the “prodrug”) to facilitatetransmittal across a cell membrane where water solubility is notbeneficial, but then it is metabolically hydrolyzed to the carboxylicacid once inside the cell where water solubility is beneficial.

Prodrugs have many useful properties. For example, a prodrug may be morewater-soluble than the ultimate drug, thereby facilitating intravenousadministration of the drug. A prodrug may also have a higher level oforal bioavailability than the ultimate drug. After administration, theprodrug is enzymatically or chemically cleaved to deliver the ultimatedrug in the blood or tissue.

Ester prodrugs of the compounds disclosed herein are specificallycontemplated. An ester may be formed from a carboxylic acid functionalgroup linked to a compound of formula (I) above by reaction with analcohol or phenol. Alternatively, an ester may be formed from a hydroxylfunctional group linked to a compound of formula (I) above by reactionwith a carboxylic acid or an aminoacid. While not intending to belimiting, an ester may be an alkyl ester, an aryl ester, or a heteroarylester. The term alkyl has the meaning generally understood by thoseskilled in the art and refers to linear, branched, or cyclic alkylmoieties. C₁₋₆alkyl esters are particularly useful, where alkyl part ofthe ester has from 1 to 6 carbon atoms and includes, but is not limitedto, methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, iso-butyl,t-butyl, pentyl isomers, hexyl isomers, cyclopropyl, cyclobutyl,cyclopentyl, cyclohexyl, and combinations thereof having from 1-6 carbonatoms.

The compounds of the present invention according to formula (I) abovecan be used for the treatment of a pathological state arising from theuncontrolled activation and/or overexpression of GSK-3β, selected fromthe group consisting of (i) insulin-resistance disorders; (ii)neurodegenerative diseases; (iii) mood disorders; (iv) schizophrenicdisorders; (v) cancerous disorders; (vi) inflammation; (vii) substanceabuse disorders; and (viii) epilepsies.

Advantageously, insulin-resistance disorders are type-2 diabetes,syndrome X, obesity and polycystic ovary syndrome.

Advantageously, acute and chronic neurodegenerative diseases areParkinson's disease, Alzheimer's disease, Huntington's disease andspinal neurodegenerative disorders.

Preferably, spinal neurodegenerative disorders are amyotrophic lateralsclerosis, multiple sclerosis, spinal muscular atrophy andneurodegeneration due to spinal cord injury.

Advantageously, mood disorders are bipolar disorders and depressivedisorders.

Preferably, bipolar disorders are bipolar I, bipolar II, cyclothymia andbipolar disorder not otherwise specified (BD-NOS),

Preferably, depressive disorders are major depressive disorder (MDD),atypical depression (AD), melancholic depression, psychotic majordepression (PMD), catatonic depression, postpartum depression (PPD),seasonal affective disorder (SAD), dysthymia, and depressive disordernot otherwise specified (DD-NOS)

Advantageously, schizophrenic disorders are paranoid schizophrenia,disorganized schizophrenia, catatonic schizophrenia, simpleschizophrenia, residual schizophrenia, and undifferentiatedschizophrenia.

Advantageously, cancerous disorders are prostate, pancreatic, ovarian,and colon-rectal cancer and MLL-associated leukaemia.

Advantageously, substance abuse disorders are abuse disorders due topsychostimulants.

Typically, the 1H-indazole-3-carboxamide according to formula (I) usefulin this invention are administered in the form of a pharmaceuticalcomposition.

Accordingly, a further aspect of the present invention relates to apharmaceutical composition comprising at least one compound of formula(I) as described above and at least one inert pharmaceuticallyacceptable excipient, for use in the treatment of a pathological statearising from the uncontrolled activation and/or over-expression ofGSK-3β, selected from the group consisting of (i) insulin-resistancedisorders, such as type-2 diabetes, syndrome X, obesity and polycysticovary syndrome; (ii) neurodegenerative diseases, such as Parkinson'sdisease, Alzheimer's disease, Huntington's disease, and spinalneurodegenerative disorders; (iii) mood disorders, such as bipolardisorders and depressive disorders; (iv) schizophrenic disorders; (v)cancerous disorders, such as prostate, pancreatic, ovarian, andcolon-rectal cancer and MLL-associated leukaemia; (vi) inflammation,(vii) substance abuse disorders; and (viii) epilepsies.

Preferably, the pharmaceutical composition of the present invention isprepared in suitable dosage forms comprising an effective amount of atleast one compound of formula (I) as described above, a salt thereofwith a pharmaceutically acceptable organic or inorganic acid or base, ora prodrug thereof, and at least one inert pharmaceutically acceptableexcipient.

Examples of suitable dosage forms are tablets, capsules, coated tablets,granules, solutions and syrups for oral administration; solutions,pomade and ointment for topical administration; medicated patches fortransdermal administration; suppositories for rectal administration andinjectable sterile solutions.

Other suitable dosage forms are those with sustained release and thosebased on liposomes for oral, injectable or transdermal administration.

The dosage forms can also contain other traditional ingredients such as:preservatives, stabilizers, surfactants, buffers, salts for regulatingosmotic pressure, emulsifiers, sweeteners, colorants, flavourings andthe like.

The amount of the 1H-indazole-3-carboxamide according to formula (I) orof the pharmaceutically acceptable salt of acid addition thereof in thepharmaceutical composition of the present invention can vary over a widerange depending on known factors, for example, the type of pathology,the severity of the disease, the patient's body weight, the dosage form,the chosen route of administration, the number of administrations perday and the efficacy of the selected 1H-indazole-3-carboxamide compoundaccording to formula (I). However, a person skilled in the art candetermine the optimum amount in easily and routinely manner.

Typically, the amount of compound of formula (I) or of thepharmaceutically acceptable salt of acid addition thereof in thepharmaceutical composition of the present invention will be such as toensure a level of administration from 0.0001 to 100 mg/kg/day.Preferably, the level of administration is from 0.001 to 50 mg/kg/day,and even more preferably from 0.01 to 10 mg/kg/day.

The dosage forms of the pharmaceutical composition of the presentinvention can be prepared by techniques that are familiar to apharmaceutical chemist, and comprise mixing, granulation, compression,dissolution, sterilization and the like.

Non-limiting examples of compounds of formula (I) that are usefulaccording to the present invention are those of the following table 1.

TABLE 1 IUPAC name Structure 1 N-({1-[2-(4-hydroxyphenyl)ethyl]-piperidin-4-yl}methyl)-5-methoxy-1H- indazole-3-carboxamide

2 5-methoxy-N-({1-[2-(4- methoxyphenyl)ethyl]piperidin-4-yl}methyl)-1H-indazole-3- carboxamide

3 N-{[1-(2,4-dichlorobenzyl)piperidin-4-yl]methyl}-5-methoxy-1H-indazole- 3-carboxamide

4 5-methoxy-N-({1-[4- (trifluoromethyl)benzyl]piepridin-4-yl}methyl)-1H-indazole-3- carboxamide

5 N-({1-[4-(benzyloxy)benzyl]piperidin-4-yl}methyl)-5-methoxy-1H-indazole- 3-carboxamide

6 N-{[1-(4-hydroxybenzyl)piperidin-4-yl]methyl}-5-methoxy-1H-indazole-3- carboxamide

EXPERIMENTAL PART

¹H-NMR spectroscopy: internal standard=Tetramethylsilane;DMSO-d₆=deuterated dimethyl sulfoxide; (s)=singlet; (d)=doublet;(t)=triplet; (br)=broad; (dd)=double doublet; (dt)=double triplet;(ddd)=double double doublet; (dtd)=double triple doublet; (m)=multiplet;J=coupling constant; δ=chemical shift (in ppm).

Preparation of Compounds of Formula (I)

Compounds of formula (I) can be obtained by methods known to personsskilled in the art, for example by the following methods A to D.

Method A

1-Hydroxybenzotriazole (HOBt, 7.40 g, 54.8 mmoles) andN,N′-dicyclohexylcarbodiimide (DCC, 11 g, 53.3 mmoles) were added to asolution of a convenient substituted 1H-indazole-3-carboxylic acid(compound i, 12 g, 49.8 mmoles) in DMF (200 ml) at 0° C. After 1 hour, asolution of a convenient 1-substituted [piperidin-4-yl]methanamine(compound ii, 10 g, 58.1 mmoles) in DMF (100 ml) was added at the sametemperature. The mixture was stirred at 0° C. for 2 hours then it wasleft to reach room temperature during the night. The mixture was dilutedwith AcOEt then the solid was removed by filtration. The solution wasextracted three time with hydrochloridric acid (HCl) 2N. The pH of theacid phase was increased (about 13) with 5N NaOH and solution wasextracted three times with dichloromethane (DCM). The organic phase wasdried with anhydrous Na₂SO₄.

The solvent was filtered, evaporated under reduced pressure and theresidue was adequately purified.

The following intermediate compounds (a-d) can be used as compound (ii)in the synthetic pathway above:

a) 1-{1-[2-(4-methoxyphenyl)ethyl]piperidin-4-yl}-methanamine

To a stirred solution ofN-[phenylmethylidene]-1-(piperidin-4-yl)methanamine (compound iii; 0.158moles; 31.9 g), prepared as described in WO2004/101548 in absoluteethanol (70 ml), 1-(2-bromoethyl)-4-methoxybenzene (compound iv; 0.237moles; 32.7 g) and potassium carbonate were added.

The solution was refluxed for 8 hours, then cooled and concentrated byevaporating the solvent under reduced pressure. The reaction mixture wasdiluted with 3N HCl and stirred at room temperature for 3 hours. Theacid solution was then washed with dichloromethane and made alkaline.The aqueous phase was extracted with three portions of dichloromethane,which were reunited and dried over Na₂SO₄.

The solvent was removed by evaporating under reduced pressure and theproduct (v) thus obtained was used as such without any furtherpurification.

¹H NMR (300 MHz, DMSO-d₆). δ 7.00-7.19 (m, 2H), 6.76-6.89 (m, 2H), 3.71(s, 3H), 2.91 (d, J=11.56 Hz, 2H), 2.55-2.72 (m, 4H), 2.37-2.47 (m, 2H),1.90 (dt, J=1.98, 11.56 Hz, 2H), 1.70 (d, J=11.89 Hz, 2H), 1.52 (ddd,J=3.96, 7.27, 10.90 Hz, 1H), 1.15 (dtd, J=3.80, 12.01, 12.14 Hz, 2H).

[M.M.+H⁺] calculated 249.1961; [M.M.+H⁺] found 249.1950.

b) 1-[1-(2,4-dichlorobenzyl)piperidin-4-yl]methanamine

The intermediate (vii) has been prepared by means of the same methoddescribed for the preparation of intermediate (v), using2,4-dichloro-1-(chloromethyl)benzene (compound vi) as starting reagent.

The product (vii) has been purified with flash chromatography (SiO₂,CHCl₃/MeOH=9/1).

c) 1-{1-[4-(trifluoromethyl)benzyl]piperidin-4-yl}methanamine

The intermediate (ix) has been prepared by means of the same methoddescribed for the preparation of intermediate (v), using1-(bromomethyl)-4-(trifluoromethyl)benzene (compound viii) as startingreagent.

The product (ix) has been purified with flash chromatography (SiO₂,CHCl₃/MeOH=9/1).

d) 1-{1-[4-(benzyloxy)benzyl]piperidin-4-yl}methanamine

The intermediate (xi) has been prepared by means of the same methoddescribed for the preparation of intermediate (v), using1-(benzyloxy)-4-(chloromethyl)benzene (compound x) as starting reagent.

The product (xi) thus obtained was used as such without any furtherpurification.

For example, compounds (5) and (6) can be prepared according to method Aas described below.

Compound (5) can be prepared using compounds (xii) and (xi) as startingmaterials, following method A disclosed above.

Compound (5) (0.6 mmoles) was hydrogenated in a micro reactor continuousflow system (H-Cube) using CartCart Pd/C 10% as cartridge. Keyparameters of H-Cube were set as follows: temperature 80° C.; pressure 1bar; flow 1 ml/minute.

The solvent was removed by evaporating under reduced pressure, and thecompound (6) was purified as disclosed in Table 2.

Method B

First Step:

To a suspension of a convenient compound (xiii) (2.13 g; 0.0061 moles)in toluene (50 ml) was added drop wise a solution of1-(1-benzylpiperidin-4-yl)methanamine (compound xiv; 2.52 g; 0.012moles), prepared as described in WO 94/10174, and triethylamine (TEA;3.2 ml; 0.023 moles) in toluene (10 ml). The reaction mixture wasrefluxed for 12 hours, and then filtered. Solvent was removed byevaporation under reduced pressure and residue was taken up with ethylacetate. The organic phase was transferred into a separated funnel,washed with saturated NaHCO₃ solution and water, separated out and driedover Na₂SO₄.

The product obtained (xv) was adequately crystallized.

Second Step:

A solution of a convenientN-[(1-benzylpiperidin-4-yl)methyl]-1H-indazole-3-carboxamide (compoundxv; 0.506 g; 1.34 mmol) in absolute ethanol (8 ml) and glacial aceticacid (0.8 ml) was hydrogenated in a micro reactor continuous flow system(H-Cube) using CartCart Pd/C 10% as cartridge. Key parameters of H-Cubewere set as follow: temperature 80°; pressure 10 bar; flow 1 ml/minute.

After three hours, the solution was concentrated by reduced pressure,diluted with water and transferred into a separating funnel. The aqueousphase was then washed with ethyl acetate, made alkaline with 1N NaOH andextracted with ethyl acetate. The organic layers were collected, driedover Na₂SO₄ and solvent was removed by evaporation under reducedpressure.

The solid thus obtained was dried in a stove under vacuum to give 0.27 gof the desired substitutedN-(piperidin-4-ylmethyl)-1H-indazole-3-carboxamide (xvi), which was usedwithout any further purification.

Third Step:

To a solution of compound (xvi) (0.75 mmol; 215 mg) inmethyl-ethyl-ketone (MEK; 9 ml) stirred at 85° C., the convenienthalogenated compound (xvii; 1.05 Eq) and triethylamine (TEA; 210 μl; 2Eq) were added drop wise. The reaction mixture was refluxed for 8 hours,then cooled and diluted with ethyl acetate. The organic layer was washedwith a saturated NH₄Cl solution and water. The organic phase wasseparated out and dried over Na₂SO₄.

The solvent was removed by evaporating under reduced pressure, and theproduct (I) was purified as described in Table 2.

Method C

First Step:

Thionyl chloride (SOCl₂; 9.3 ml; 0.128 moles) was added to a suspensionof a convenient substituted 1H-indazole-3-carboxylic acid (compound i;2.36 g; 0.0123 moles) in toluene (77 ml), and the reaction mixture wasrefluxed for 4 hours. The solvent was removed by evaporation underreduced pressure and the residue was taken up twice in toluene to give2.13 g of the desired product (xiii) 2,10-substituted7H,14H-pyrazino[1,2-b:4,5-b′]di-indazole-7,14-dione.

Second Step:

To a suspension of (xiii) (5.2 mmol) in toluene (40 ml), a solution ofthe convenient amine (compound ii; 2.1 Eq) and triethylamine (TEA; 3.6Eq; 2.6 ml) was added drop wise. The mixture reaction was refluxed for 8hours, then cooled and stirred in 2N HCl (20 ml) for 8 hours. Thesuspension was transferred in a separating funnel and aqueous phase wasseparating out and made alkaline with 1N NaOH.

The solvent was removed by evaporating under reduced pressure, and theproduct (I) was purified as described below.

For example, compound (3) can be prepared following method C describedbelow.

Thionyl chloride (SOCl₂; 9.3 ml; 0.128 moles) was added to a suspensionof 5-methoxy-1H-indazole-3-carboxylic acid (compound xii; 2.36 g; 0.0123moles) in toluene (77 ml), and the reaction mixture was refluxed for 4hours. The solvent was removed by evaporation under reduced pressure andthe residue was taken up twice in toluene to give 2.13 g of the desiredproduct2,10-dimethoxy-7H,14H-pyrazino[1,2-b:4,5-b′]di-indazole-7,14-dione(xviii).

¹H NMR (300 MHz, CHLOROFORM-d): δ 8.53 (dd, J=0.58, 9.17 Hz, 2H), 7.64(d, J=1.98 Hz, 2H), 7.35 (dd, J=2.48, 9.08 Hz, 2H), 3.97 (s, 6H).

[M.M.+H⁺] calculated 349.0937; [M.M.+H⁺] found 349.0922.

To a suspension of compound (xviii) (2.13 g; 0.0061 moles) in toluene(50 ml) was added drop wise a solution of the intermediate compound(vii) (2.52 g; 0.012 moles), prepared as described in method A, andtriethylamine (TEA; 3.2 ml; 0.023 moles) in toluene (10 ml). Thereaction mixture was refluxed for 12 hours, and then filtered. Solventwas removed by evaporation under reduced pressure and residue was takenup with ethyl acetate. The organic phase was transferred into aseparated funnel, washed with saturated NaHCO₃ solution and water,separated out and dried over Na₂SO₄.

Compound (3) thus obtained was crystallized as disclosed in followingTable 2.

¹H NMR (300 MHz, DMSO-d₆): δ 13.39 (br. s., 1H), 8.25 (t, J=6.04 Hz,1H), 7.56 (d, J=2.01 Hz, 1H), 7.50 (dd, J=0.55, 8.96 Hz, 1H), 7.17-7.36(m, 5H), 7.05 (dd, J=2.47, 9.06 Hz, 1H), 3.80 (s, 3H), 3.43 (s, 2H),3.20 (t, J=6.13 Hz, 2H), 2.79 (d, J=11.16 Hz, 2H), 1.89 (t, J=10.61 Hz,2H), 1.46-1.74 (m, 3H), 1.07-1.34 (m, 2H).

[M.M.+H⁺] calculated 379.2134; [M.M.+H⁺] found 379.2129.

Method D

A solution of product (xix), a conveniently substituted arylboronic acid(compound xx),[1,1′-bis(diphenylphosphino)ferrocene]-dichloro-palladium(11)[Pd(dppf)Cl₂], caesium carbonate in 1,4-dioxane and water (ratio 3:1)was subjected to microwave irradiation.

Programme was set as follows:

-   -   3′; T₁=160° C., T₂=130° C.; max power 300 W    -   45′; T₁=160° C., T₂=130° C.; max power 300 W    -   5′; T₁=20° C., T₂=15° C.

After one cycle of microwave irradiation, solvents were removed byevaporating under reduce pressure and the reaction mixture was dilutedwith a solution of chloroform and methanol in a 2:1 ratio and filtered.

Products (I) thus obtained were purified as described below.

Purification Methods

Compounds of formula (I), obtained according to one of method A to D,can be purified with one of the following techniques (a)-(c).

(a) Flash Chromatography on Silica Gel.

Flash chromatography was carried out with a Biotage Flash MasterPersonal system on 20-45 μm silica cartridge or Grace Reveleris flashchromatography system with 40 μM silica cartridge.

Flow=60 ml/min.

The solvents used as eluents are shown in the following Table 2.

(b) Crystallization

A different crystallization solvent was used depending on the compoundto be purified. The solvents are shown in the following Table 2.

(c) Preparative LC/MS System.

LC/MS system consisted of a Waters 2767 Sample manager, a Waters 2478dual λ absorbance detector and a Waters Micromass ZQ single quadrupolemass spectrometer with an electrospray ionization (ESI) source. Thecolumn used was a X-Bridge Prep C18 5 μm with 19×10 mm (Waters)pre-column. Fraction collection was available from the system softwareMassLynx™ v. 4.1. Detection wavelength was set to 230 nm and temperatureto 25° C.

The sample was dissolved (50 mg/ml) in DMSO/CH₃CN in 1:1 ratio. Themobile phase was:channel A=CH₃CN+0.1% formic acid (Eluent A)channel B=H₂O+0.1% formic acid (Eluent B)

-   -   flow=40 ml/min    -   gradient=minimum and maximum percentage of eluent A reached in        15 minutes ranges from 2 to 20 and from 25 to 55, respectively.

The following Table 2 shows both the preparation and the purificationmethod for each compound of formula (I), as listed in Table 1, and themonoisotopic mass for each compound.

TABLE 2 Parameters Prep- or solvent MM MM aration Purification used forthe founded calculated N^(o) method method purification [M + H⁺] [M +H⁺] 1 B (b) CHCl₃ 409.2226 409.2234 2 C (b) EtOH abs/AcOEt 423.2402423.2396 3 C (b) Hex/AcOEt 447.1357 447.1349 4 C (a) CHCl₃/MeOH 447.2003447.2002 5 A or C (b) THF 485.2548 485.2547 6 A (a) CHCl₃/MeOH 395.2083395.2078 MM: monoisotopic mass CHCl₃: chloroform EtOH abs: absoluteethanol AcOEt: ethyl acetate Hex: hexane MeOH: methanol THF:tetrahydrofurane

TABLE 3 N^(o) 1H-NMR peaks 1 DMSO-d6; δ 13.40 (br. S., 1H), 9.07 (s,1H), 8.26 (t, J = 6.16 Hz, 1H), 7.55 (d, J = 2.42 Hz, 1H), 7.50 (dd, J =0.61, 9.08 Hz, 1H), 7.05 (dd, J = 2.50, 9.10 Hz, 1H), 6.94-7.02 (m, 2H),6.60-6.68 (m, 2H), 3.80 (s, 3H), 3.19 (t, J = 6.36 Hz, 2H), 2.89 (d, J =11.30 Hz, 2H), 2.53-2.65 (m, 2H), 2.30-2.46 (m, 2H), 1.79-2.00 (m, 2H),1.47-1.74 (m, 3H), 1.06-1.31 (m, 2H) 2 DMSO-d6; δ 13.62 (br. s., 1H),8.24 (t, J = 6.06 Hz, 1H), 7.55 (d, J = 2.42 Hz, 1H), 7.52 (dd, J =0.61, 8.88 Hz, 1H), 7.07-7.16 (m, 2H), 7.03 (dd, J = 2.42, 8.88 Hz, 1H),6.75-6.87 (m, 2H), 3.80 (s, 3H), 3.71 (s, 3H), 3.19 (t, J = 6.26 Hz,2H), 2.90 (d, J = 11.10 Hz, 2H), 2.57-2.74 (m, 2H), 2.34-2.47 (m, 2H),1.80-1.98 (m, 2H), 1.46-1.76 (m, 3H), 1.08-1.30 (m, 2H) 3 DMSO-d6; δ13.38 (br. s., 1H), 8.28 (t, J = 6.22 Hz, 1H), 7.56 (t, J = 2.38 Hz,2H), 7.52 (d, J = 2.56 Hz, 1H), 7.50 (d, J = 1.83 Hz, 1H), 7.41 (dd, J =2.20, 8.20 Hz, 1H), 7.06 (dd, J = 2.56, 9.15 Hz, 1H), 3.81 (s, 3H), 3.51(s, 2H), 3.21 (t, J = 6.22 Hz, 2H), 2.80 (d, J = 11.34 Hz, 2H), 2.01 (t,J = 10.79 Hz, 2H), 1.48-1.78 (m, 3H), 1.07-1.35 (m, 2H) 4 DMSO-d₆; δ13.40 (s, 1H), 8.27 (t, J = 6.10 Hz, 1H), 7.67 (d, J = 8.01 Hz, 2H),7.47-7.57 (m, 4H), 7.05 (dd, J = 2.44, 9.06 Hz, 1H), 3.80 (s, 3H), 3.53(s, 2H), 3.20 (t, J = 6.27 Hz, 2H), 2.78 (d, J = 11.50 Hz, 2H), 1.94 (t,J = 10.80 Hz, 2H), 1.47-1.77 (m, 3H), 1.12-1.36 (m, 2H) 5 DMSO-d₆; δ13.44 (br. s., 1H), 8.25 (t, J = 6.22 Hz, 1H), 7.55 (d, J = 1.83 Hz,1H), 7.50 (d, J = 9.15 Hz, 1H), 7.28-7.46 (m, 5H), 7.14-7.21 (m, 2H),7.04 (dd, J = 2.38, 8.96 Hz, 1H), 6.88-6.98 (m, 2H), 5.07 (s, 2H), 3.79(s, 3H), 3.35 (s, 2H), 3.18 (t, J = 6.22 Hz, 2H), 2.77 (d, J = 11.34 Hz,2H), 1.85 (t, J = 10.79 Hz, 2H), 1.45-1.71 (m, 3H), 1.00-1.32 (m, 2H) 6DMSO-d₆; δ 13.50 (s, 1H), 9.70 (br. s., 1H), 8.35 (d, J = 17.17 Hz, 1H),7.55 (d, J = 2.31 Hz, 1H), 7.51 (d, J = 8.92 Hz, 1H), 7.27 (br. s., 2H),7.05 (dd, J = 2.31, 8.92 Hz, 1H), 6.78 (d, J = 7.60 Hz, 2H), 3.80 (s,3H), 2.60-3.59 (m, 8H), 0.99-2.15 (m, 5H) DMSO: dimethyl sulfoxide

The compounds 7 to 31 were prepared as described hereinbelow.

Synthesis of compound 7—Methyl2-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-4-carboxylate7a) Tert-butyl4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidine-1-carboxylate

1-Hydroxybenzotriazole (HOBt, 24.3 g, 142 mmoles) andN,N′-dicyclohexylcarbodiimide (DCC, 29.3 g, 142 mmoles) were added to asolution of 5-methoxy-1H-indazole-3-carboxylic acid (30 g, 129 mmoles)in DMF (400 mL) at 0° C. After 1 hour, a solution ofethyl[4-(aminomethyl)piperidin-1-yl]acetate (26 g, 129 mmoles) in DMF(250 mL) was added at the same temperature. The mixture was stirred at0° C. for 2 hours then was left to reach room temperature during thenight. The mixture was diluted with EtOAc and the solid was removed byfiltration. The solution was extracted three times with hydrochloridricacid (HCl) 2N. The pH of the acid phase was increased (about 13) with 5NNaOH and the solution was extracted three times with dichloromethane(DCM). The organic phase was dried over anhydrous Na₂SO₄ and the solventwas filtered and evaporated under reduced pressure providing Tert-butyl4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidine-1-carboxylate7a (96% yield).

MS: 389 m/z (M+H)⁺.

7b) 5-Methoxy-N-(piperidin-4-ylmethyl)-1H-indazole-3-carboxamidehydrochloride

2 M HCl in Et₂O (1.8 L) was added to a solution of compound 7a (92.8 g,0.24 moles) in MeOH (500 mL). The mixture was stirred for 3 hours atroom temperature then the resulting solid was filtered and dried to give5-methoxy-N-(piperidin-4-ylmethyl)-1H-indazole-3-carboxamidehydrochloride 7b (61.1 g, 89% yield).

MS: 289 m/z (M+H)⁺.

Finally, a mixture of compound 7b (637 mg, 1.96 mmoles) and potassiumcarbonate (813 mg, 5.88 mmoles) in acetonitrile (5 mL) was heated toreflux for 1 hour, then a solution of methyl2-(chloromethyl)-1,3-thiazole-4-carboxylate (500 mg, 2.6 mmoles) inacetonitrile (5 mL) was added dropwise. The mixture was refluxedovernight then was cooled, diluted with EtOAc and filtered. Theresulting solid was washed with water, dried and purified via flashchromatography (silica, from CHCl₃ to CHCl₃:MeOH 9:1) providing 280 mg(32% yield) of methyl2-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-4-carboxylate7.

¹H NMR (300 MHz, DMSO-d6) δ=13.41 (s, 1H), 8.46 (s, 1H), 8.29 (t, J=6.0Hz, 1H), 7.55 (d, J=2.2 Hz, 1H), 7.50 (d, J=9.1 Hz, 1H), 7.05 (dd,J=2.4, 9.0 Hz, 1H), 3.81 (s, 3H), 3.80 (s, 5H), 3.21 (t, J=6.2 Hz, 2H),2.89 (d, J=11.3 Hz, 2H), 2.13 (t, J=10.8 Hz, 2H), 1.78-1.54 (m, 3H),1.37-1.14 (m, 2H)

MS: 444 m/z (M+H)⁺.

Synthesis of compound8—2-{[4-({[(5-Methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-4-carboxylicacid

To a solution of compound 7 (1.85 mmoles) in MeOH (10 mL) aqueous 1MNaOH (3.7 mL) was added. The solution was refluxed overnight then theorganic solvent was removed under vacuum, the residue was diluted withH₂O and the pH was adjusted to 5 by adding 1M HCl. The mixture was keptat 4° C. overnight then the resulting solid was filtered, washed withfresh water and dried under vacuum to give2-{[4-({[(5-Methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-4-carboxylicacid 8 (43% yield).

¹H NMR (300 MHz, DMSO-d6) δ=13.42 (br. s., 1H), 12.91 (br. s., 1H), 8.34(s, 1H), 8.29 (t, J=6.0 Hz, 1H), 7.56 (d, J=2.2 Hz, 1H), 7.51 (d, J=9.1Hz, 1H), 7.05 (dd, J=2.6, 9.1 Hz, 1H), 3.87-3.69 (m, 5H), 3.22 (t, J=6.2Hz, 2H), 2.89 (d, J=11.3 Hz, 2H), 2.12 (t, J=10.6 Hz, 2H), 1.81-1.50 (m,3H), 1.37-1.11 (m, 2H)

MS: 430 m/z (M+H)⁺.

Synthesis of compound 9—Methyl2-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-oxazole-4-carboxylate

Methyl 2-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-oxazole-4-carboxylate 9 was prepared,according to the procedure described for compound 7, using methyl2-(chloromethyl)-1,3-oxazole-4-carboxylate. Yield: 410 mg, 45%.

¹H NMR (300 MHz, DMSO-d6) δ=13.40 (br. s., 1H), 8.80 (s, 1H), 8.26 (t,J=6.2 Hz, 1H), 7.55 (d, J=2.6 Hz, 1H), 7.51 (d, J=9.1 Hz, 1H), 7.05 (dd,J=2.4, 9.0 Hz, 1H), 3.80 (s, 6H), 3.67 (s, 2H), 3.18 (t, J=6.2 Hz, 2H),2.82 (d, J=11.3 Hz, 2H), 2.14-1.93 (m, 2H), 1.74-1.45 (m, 3H), 1.29-1.10(m, 2H)

MS: 428 m/z (M+H)⁺.

Synthesis of compound10—2-{[4-({[(5-Methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-oxazole-4-carboxylicacid hydrate

2-{[4-({[(5-Methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-oxazole-4-carboxylic acid hydrate 10 wasprepared, according to the procedure described for compound 8, startingfrom compound 9. Yield: 238 mg, 82%.

¹H NMR (300 MHz, DMSO-d6) δ=13.40 (s, 1H), 12.99 (br. s., 1H), 8.67 (s,1H), 8.26 (t, J=6.0 Hz, 1H), 7.55 (d, J=2.2 Hz, 1H), 7.50 (d, J=9.5 Hz,1H), 7.04 (dd, J=2.6, 9.1 Hz, 1H), 3.80 (s, 3H), 3.66 (s, 2H), 3.18 (t,J=6.4 Hz, 2H), 2.82 (d, J=11.0 Hz, 2H), 2.05 (t, J=10.4 Hz, 2H),1.76-1.44 (m, 3H), 1.33-1.05 (m, 2H)

MS: 414 m/z (M+H)⁺.

Synthesis of compound 11—Methyl2-{[4-({[(5-bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-oxazole-4-carboxylate11a) Tert-butyl4-({[(5-bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidine-1-carboxylate

Tert-butyl4-({[(5-bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidine-1-carboxylate11a was prepared, according to the procedure described for compound 7,from 5-bromo-1H-indazole-3-carboxylic acid and tert-butyl4-(aminomethyl)piperidine-1-carboxylate. Yield: 40.6 g, 87%

MS: 437 m/z (M+H)⁺.

11b) 5-Bromo-N-(piperidin-4-ylmethyl)-1H-indazole-3-carboxamidehydrochloride

2M HCl in Et₂O (1.8 L) was added to a solution of compound tert-butyl4-(aminomethyl)piperidine-1-carboxylate 11a (0.24 moles) in MeOH (500mL). The mixture was stirred for 3 hours at room temperature then theresulting solid was filtered and dried to give5-Bromo-N-(piperidin-4-ylmethyl)-1H-indazole-3-carboxamide hydrochloride11 b (76% yield).

MS: 337 m/z (M+H)⁺.

Finally, methyl2-{[4-({[(5-bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-oxazole-4-carboxylate11 was prepared, according to the procedure described for compound 7,from 11 b and methyl 2-(chloromethyl)-1,3-oxazole-4-carboxylate. Yield:166 mg, 16%.

¹H NMR (300 MHz, DMSO-d6) δ=13.73 (br. s., 1H), 8.80 (s, 1H), 8.42 (t,J=6.0 Hz, 1H), 8.31 (dd, J=0.8, 1.8 Hz, 1H), 7.60 (dd, J=0.8, 8.8 Hz,1H), 7.52 (dd, J=1.8, 8.8 Hz, 1H), 3.80 (s, 3H), 3.67 (s, 2H), 3.18 (t,J=6.4 Hz, 2H), 2.81 (d, J=11.3 Hz, 2H), 2.13-1.95 (m, 2H), 1.74-1.44 (m,3H), 1.32-1.06 (m, 2H)

MS: 428 m/z (M+H)⁺.

Synthesis of compound12—2-({4-[({[5-(2,3-Difluorophenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)-1,3-oxazole-4-carboxylicacid hydrate

A solution of compound 11 (0.44 mmoles), (2,3-difluorophenyl)boronicacid (1.77 mmoles),[1,1′-bis(diphenylphosphino)ferrocene]-dichloro-palladium(II)[Pd(dppf)Cl₂] (81 mg, 0.11 mmoles) and caesium carbonate (575 mg, 1.76mmoles) in 1,4-dioxane and water (ratio 3:1, 8 mL) was subjected tomicrowave irradiation as follows:

-   -   Time period=3′; T₁=160° C., T₂=130° C.; max power 300 W    -   Time period=45′; T₁=160° C., T₂=130° C.; max power 300 W    -   Time period=5′; T₁=20° C., T₂=15° C.

After one cycle of microwave irradiation, solvents were removed byevaporating under reduce pressure and the reaction mixture was dilutedwith a solution of methanol (20 mL), filtered over Celite and driedunder vacuum. The crude product was filtered on a silica cartridge andwashed with chloroform and methanol in a 1:1 ratio. The resulting solidwas dissolved in DMSO and purified via preparative HPLC (channelA=CH₃CN+0.1% formic acid; channel B=H₂O+0.1% formic acid: flow=40ml/min; gradient=15%-50% of eluent A in 15 minutes), providing2-({4-[({[5-(2,3-Difluorophenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)-1,3-oxazole-4-carboxylicacid hydrate 12 (6% yield).

¹H NMR (300 MHz, DMSO-d6) δ=13.70 (s, 1H), 12.99 (br. s., 1H), 8.57 (s,1H), 8.42 (t, J=6.0 Hz, 1H), 8.34 (d, J=0.7 Hz, 1H), 7.73 (dd, J=0.8,8.8 Hz, 1H), 7.61 (td, J=1.8, 8.7 Hz, 1H), 7.52-7.21 (m, 3H), 3.64 (s,2H), 3.20 (t, J=6.2 Hz, 2H), 2.82 (d, J=11.0 Hz, 2H), 2.04 (t, J=10.6Hz, 2H), 1.73-1.45 (m, 3H), 1.33-1.09 (m, 2H)

MS: 496 m/z (M+H)⁺.

Synthesis of compound 13—Ethyl4-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-2-carboxylate

Ethyl 4-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-2-carboxylate 13 was prepared,according to the procedure described for compound 7, using ethyl4-(chloromethyl)-1,3-thiazole-2-carboxylate. Yield: 45 mg, 11%.

¹H NMR (300 MHz, DMSO-d6) δ=13.39 (s, 1H), 8.26 (t, J=6.0 Hz, 1H), 7.86(s, 1H), 7.55 (d, J=2.2 Hz, 1H), 7.50 (d, J=8.8 Hz, 1H), 7.04 (dd,J=2.2, 8.8 Hz, 1H), 4.37 (q, J=7.0 Hz, 2H), 3.80 (s, 3H), 3.64 (s, 2H),3.19 (t, J=6.2 Hz, 2H), 2.85 (d, J=11.3 Hz, 2H), 1.98 (t, J=10.6 Hz,2H), 1.79-1.45 (m, 3H), 1.33 (t, J=7.1 Hz, 3H), 1.29-0.96 (m, 2H)

MS: 458 m/z (M+H)⁺.

Synthesis of compound 14—Methyl2-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-3-carboxylate

Methyl 2-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-3-carboxylate 14 was prepared, according tothe procedure described for compound 7, using methyl2-(chloromethyl)furan-3-carboxylate. Yield: 120 mg, 13%.

¹H NMR (300 MHz, DMSO-d6) δ=13.39 (s, 1H), 8.23 (t, J=6.0 Hz, 1H), 7.70(d, J=2.2 Hz, 1H), 7.54 (d, J=2.6 Hz, 1H), 7.50 (d, J=8.4 Hz, 1H), 7.04(dd, J=2.4, 9.0 Hz, 1H), 6.70 (d, J=2.2 Hz, 1H), 3.83 (s, 2H), 3.80 (s,3H), 3.76 (s, 3H), 3.17 (t, J=6.4 Hz, 2H), 2.80 (d, J=11.3 Hz, 2H),2.10-1.88 (m, 2H), 1.70-1.42 (m, 3H), 1.31-1.02 (m, 2H)

MS: 427 m/z (M+H)⁺.

Synthesis of compound 15—Ethyl5-{[4-({[(5-bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylate

Ethyl 5-{[4-({[(5-bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylate 15 was prepared, according tothe procedure described for compound 7, from 11b and ethyl5-(chloromethyl)furan-2-carboxylate. Yield: 300 mg, 62%.

¹H NMR (300 MHz, DMSO-d6) δ=13.73 (br. s., 1H), 8.41 (t, J=6.04 Hz, 1H),8.32 (dd, J=0.73, 1.83 Hz, 1H), 7.57-7.65 (m, 1H), 7.45-7.56 (m, 1H),7.21 (d, J=3.66 Hz, 1H), 6.48 (d, J=3.66 Hz, 1H), 4.27 (q, J=7.32 Hz,2H), 3.53 (s, 2H), 3.19 (t, J=6.40 Hz, 2H), 2.81 (d, J=11.34 Hz, 2H),1.82-2.09 (m, 2H), 1.64 (d, J=12.44 Hz, 3H), 1.02-1.36 (m, 5H)

MS: 489 m/z (M+H)⁺.

Synthesis of compound16—5-({4-[({[5-(2-Methoxypyridin-3-yl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylic acid hydrate

5-({4-[({[5-(2-Methoxypyridin-3-yl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid hydrate 16 was prepared, according to the procedure described forcompound 12, from compound 15 and (2-methoxypyridin-3-yl)boronic acidand using the following preparative HPLC parameters for thepurification: channel A=CH₃CN+0.1% formic acid; channel B=H₂O+0.1%formic acid: flow=40 ml/min; gradient=10%-45% of eluent A in 15 minutes.Yield: 14 mg, 5%.

¹H NMR (300 MHz, DMSO-d6) δ 13.62 (br. s., 1H), 8.36 (t, J=6.04 Hz, 1H),8.28 (s, 1H), 8.19 (dd, J=1.83, 5.12 Hz, 1H), 7.77 (dd, J=2.20, 7.32 Hz,1H), 7.65 (dd, J=0.80, 8.80 Hz, 1H), 7.58 (dd, J=1.80, 8.80 Hz, 1H),7.11 (dd, J=5.12, 7.32 Hz, 1H), 6.84 (br. s., 1H), 6.31 (d, J=2.93 Hz,1H), 3.89 (s, 3H), 3.47 (s, 2H), 3.19 (t, J=6.22 Hz, 2H), 2.99 (s, 1H),2.82 (d, J=10.98 Hz, 2H), 1.83-2.04 (m, 2H), 1.41-1.75 (m, 3H),1.06-1.34 (m, 2H)

MS: 490 m/z (M+H)⁺.

Synthesis of compound17—5-({4-[({[5-(6-Methoxypyridin-3-yl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)uran-2-carboxylicacid hydrate

5-({4-[({[5-(6-Methoxypyridin-3-yl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid hydrate 17 was prepared, according to the procedure described forcompound 12, from compound 15 and (6-methoxypyridin-3-yl)boronic acidand using the following preparative HPLC parameters for thepurification: channel A=CH₃CN+0.1% formic acid; channel B=H₂O+0.1%formic acid: flow=40 ml/min; gradient=10%-45% of eluent A in 15 minutes.Yield: 23 mg, 8%.

¹H NMR (300 MHz, DMSO-d6) δ 13.65 (br. s., 1H), 8.48 (d, J=2.02 Hz, 1H),8.39 (t, J=6.06 Hz, 1H), 8.34 (s, 1H), 7.96-8.07 (m, 1H), 7.70 (d,J=1.21 Hz, 2H), 7.03 (d, J=3.23 Hz, 1H), 6.94 (d, J=8.07 Hz, 1H), 6.40(d, J=3.23 Hz, 1H), 3.92 (s, 3H), 3.51 (s, 2H), 3.21 (t, J=6.26 Hz, 2H),2.83 (d, J=10.90 Hz, 2H), 1.98 (t, J=10.90 Hz, 2H), 1.48-1.78 (m, 3H),1.07-1.34 (m, 2H)

MS: 490 m/z (M+H)⁺.

Synthesis of compound18—5-({4-[({[5-(4-Methoxyphenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid hydrate

5-({4-[({[5-(4-Methoxyphenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid hydrate 18 was prepared, according to the procedure described forcompound 12, from compound 15 and (4-methoxyphenyl)boronic acid andusing the following preparative HPLC parameters for the purification:channel A=CH₃CN+0.1% formic acid; channel B=H₂O+0.1% formic acid:flow=40 ml/min; gradient=15%-50% of eluent A in 15 minutes. Yield: 14mg, 5%.

¹H NMR (300 MHz, DMSO-d6) δ 13.55 (s, 1H), 8.27-8.41 (m, 2H), 7.64-7.72(m, 2H), 7.61 (d, J=8.88 Hz, 2H), 7.05 (d, J=8.88 Hz, 2H), 6.96 (br. s.,1H), 6.37 (d, J=3.23 Hz, 1H), 3.81 (s, 3H), 3.49 (s, 2H), 3.20 (t,J=6.26 Hz, 2H), 2.82 (d, J=10.90 Hz, 2H), 1.86-2.05 (m, 2H), 1.66 (d,J=12.11 Hz, 3H), 1.09-1.33 (m, 2H)

MS: 489 m/z (M+H)⁺.

Synthesis of compound19—5-({4-[({[5-(2,3-Difluorophenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid

5-({4-[({[5-(2,3-Difluorophenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid 19 was prepared, according to the procedure described for compound12, from compound 15 and (2,3-difluorophenyl)boronic acid and using thefollowing preparative HPLC parameters for the purification: channelA=CH₃CN+0.1% formic acid; channel B=H₂O+0.1% formic acid: flow=40ml/min; gradient=15%-50% of eluent A in 15 minutes. Yield: 32 mg, 11%.

¹H NMR (300 MHz, DMSO-d6) δ 13.74 (br. s., 1H), 8.42 (t, J=5.65 Hz, 1H),8.35 (s, 1H), 7.69-7.80 (m, 1H), 7.55-7.67 (m, 1H), 7.21-7.54 (m, 3H),7.05 (d, J=3.23 Hz, 1H), 6.41 (d, J=3.23 Hz, 1H), 3.52 (s, 2H), 3.20 (t,J=6.06 Hz, 2H), 2.83 (d, J=10.50 Hz, 2H), 1.98 (t, J=10.70 Hz, 2H),1.42-1.79 (m, 3H), 1.04-1.35 (m, 2H)

MS: 495 m/z (M+H)⁺.

Synthesis of compound20—5-({4-[({[5-(2-Fluorophenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid hydrate

5-({4-[({[5-(2-Fluorophenyl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid hydrate 20 was prepared, according to the procedure described forcompound 12, from compound 15 and (2-fluorophenyl)boronic acid and usingthe following preparative HPLC parameters for the purification: channelA=CH₃CN+0.1% formic acid; channel B=H₂O+0.1% formic acid: flow=40ml/min; gradient=10%-45% of eluent A in 15 minutes. Yield: 20 mg, 7%.

¹H NMR (300 MHz, DMSO-d6) δ 13.66 (br. s., 1H), 8.39 (t, J=6.04 Hz, 1H),8.32 (s, 1H), 7.65-7.76 (m, 1H), 7.50-7.63 (m, 2H), 7.38-7.50 (m, 1H),7.23-7.38 (m, 2H), 7.08 (d, J=3.29 Hz, 1H), 6.43 (d, J=3.29 Hz, 1H),3.52 (s, 2H), 3.20 (t, J=6.22 Hz, 2H), 2.82 (d, J=10.98 Hz, 2H), 1.98(t, J=10.79 Hz, 2H), 1.44-1.79 (m, 3H), 1.02-1.38 (m, 2H)

MS: 477 m/z (M+H)⁺.

Synthesis of compound21—5-({4-[({[5-(4-Methoxypyridin-3-yl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylic acid formate

5-({4-[({[5-(4-Methoxypyridin-3-yl)-1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan-2-carboxylicacid formate 21 was prepared, according to the procedure described forcompound 12, from compound 15 and (4-methoxypyridin-3-yl)boronic acidand using the following preparative HPLC parameters for thepurification: channel A=CH₃CN+0.1% formic acid; channel B=H₂O+0.1%formic acid: flow=40 ml/min; gradient=2%-40% of eluent A in 15 minutes.Yield: 40 mg, 14%.

¹H NMR (300 MHz, DMSO-d6) δ 13.61 (br. s., 1H), 8.47 (d, J=5.85 Hz, 1H),8.31-8.43 (m, 2H), 8.24 (s, 1H), 7.66 (d, J=8.78 Hz, 1H), 7.53 (dd,J=1.46, 8.42 Hz, 1H), 7.18 (d, J=5.49 Hz, 1H), 7.09 (d, J=3.29 Hz, 1H),6.43 (d, J=3.29 Hz, 1H), 3.86 (s, 3H), 3.54 (s, 2H), 3.19 (t, J=6.04 Hz,2H), 2.83 (d, J=10.98 Hz, 2H), 1.99 (t, J=10.79 Hz, 2H), 1.44-1.79 (m,3H), 0.98-1.36 (m, 2H)

MS: 490 m/z (M+H)⁺.

Synthesis of compound 22—5-{[4-({[(5-Bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylic acid

5-{[4-({[(5-Bromo-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylicacid 22 was prepared, according to the procedure described for compound8, starting from compound 15 and using EtOH as solvent. Yield: 264 mg,98%.

¹H NMR (300 MHz, DMSO-d6) δ 13.78 (br. s., 1H), 8.43 (t, J=5.85 Hz, 1H),8.32 (d, J=1.21 Hz, 1H), 7.61 (d, J=8.80 Hz, 1H), 7.53 (dd, J=2.00, 8.80Hz, 1H), 7.11 (d, J=3.63 Hz, 1H), 6.45 (d, J=3.23 Hz, 1H), 3.57 (s, 2H),3.19 (t, J=6.26 Hz, 2H), 2.85 (d, J=11.30 Hz, 2H), 2.02 (t, J=10.90 Hz,2H), 1.45-1.77 (m, 3H), 1.08-1.37 (m, 2H)

MS: 461 m/z (M+H)⁺.

Synthesis of compound 23—Ethyl5-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylate

Ethyl 5-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylate 23 was prepared, according tothe procedure described for compound 7, starting from ethyl5-(chloromethyl)furan-2-carboxylate. Yield: 290 mg, 71%.

¹H NMR (300 MHz, DMSO-d6) δ 13.37 (br. s., 1H), 8.25 (t, J=6.04 Hz, 1H),7.42-7.60 (m, 2H), 7.21 (d, J=3.29 Hz, 1H), 6.97-7.12 (m, 1H), 6.48 (d,J=3.29 Hz, 1H), 4.26 (q, J=7.32 Hz, 2H), 3.80 (s, 3H), 3.53 (s, 2H),3.18 (t, J=6.22 Hz, 2H), 2.81 (d, J=11.34 Hz, 2H), 1.87-2.05 (m, 2H),1.46-1.73 (m, 3H), 1.28 (t, J=6.95 Hz, 3H), 1.01-1.41 (m, 2H)

MS: 441 m/z (M+H)⁺.

Synthesis of compound24—5-{[4-({[(5-Methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylicacid

5-{[4-({[(5-Methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylic acid 24 was prepared, accordingto the procedure described for compound 8, starting from compound 23 andusing EtOH as solvent. Yield: 64 mg, 84%.

¹H NMR (300 MHz, DMSO-d6) δ 12.78-14.43 (m, 1H), 8.26 (t, J=6.04 Hz,1H), 7.55 (d, J=2.56 Hz, 1H), 7.52 (d, J=9.15 Hz, 1H), 7.04 (dd, J=2.60,9.10 Hz, 1H), 6.91 (d, J=3.29 Hz, 1H), 6.35 (d, J=3.29 Hz, 1H), 4.04(br. s., 1H), 3.80 (s, 3H), 3.50 (s, 2H), 3.18 (t, J=6.22 Hz, 2H), 2.83(d, J=11.34 Hz, 2H), 1.97 (t, J=10.79 Hz, 2H), 1.47-1.73 (m, 3H),1.04-1.33 (m, 2H)

MS: 413 m/z (M+H)⁺.

Synthesis of compound 25—N-[(1-{2-[(2R,6S)-2,6-dimethylmorpholin-4-yl]ethyl}piperidin-4-yl)methyl]-5-methoxy-1H-indazole-3-carboxamide

A mixture of compound 7b (8 g, 24.6 mmoles) and potassium carbonate (17g, 123 mmoles) in acetone (250 mL) was refluxed for 1 hour, then(2R,6S)-4-(2-chloroethyl)-2,6-di methyl morpholine (25.9 mmoles) wasadded dropwise. The mixture was refluxed overnight then was cooled andfiltered. The resulting solid was dried and purified via preparativeHPLC (channel A=CH₃CN+0.1% formic acid; channel B=H₂O+0.1% formic acid:flow=40 ml/min; gradient=10%-45% of eluent A in 15 minutes) providingN-[(1-{2-[(2R,6S)-2,6-dimethylmorpholin-4-yl]ethyl}piperidin-4-yl)methyl]-5-methoxy-1H-indazole-3-carboxamide25 (48.3% yield)

¹H NMR (300 MHz, DMSO-d6) δ=13.40 (s, 1H), 8.30-8.14 (t, J=6.11 Hz, 1H),7.58-7.53 (d, J=1.98 Hz, 1H), 7.53-7.46 (dd, J=8.92, 0.66 Hz, 1H),7.11-6.96 (dd, J=8.92, 2.31 Hz, 1H), 3.80 (s, 3H), 3.57-3.43 (m, 2H),3.21-3.11 (t, J=6.28 Hz, 2H), 2.92-2.77 (d, J=11.23 Hz, 2H), 2.76-2.63(d, J=10.24 Hz, 2H), 2.44-2.26 (m, 4H), 1.97-1.77 (t, J=10, 90 Hz, 2H),1.71-1.46 (t, J=10.73 Hz, 4H), 1.27-1.07 (m, 3H), 1.06-0.94 (d, J=6.28Hz, 6H)

LC-MS: 430.28 (MH+)

Synthesis of compound 26—N-[(1-{3-[(2R,6S)-2,6-dimethylmorpholin-4-yl]propyl}piperidin-4-yl)methyl]-5-methoxy-1H-indazole-3-carboxamide

N-[(1-{3-[(2R,6S)-2,6-dimethylmorpholin-4-yl]propyl}piperidin-4-yl)methyl]-5-methoxy-1H-indazole-3-carboxamide26, was prepared, according to the procedure described for compound 25,using (2R,6S)-4-(3-chloropropyl)-2,6-dimethylmorpholine and methanol assolvent. Yield=91 mg (59.1%).

¹H NMR (300 MHz, DMSO-d6) δ=12.12 (s, 1H), 7.80-7.62 (d, J=2.20 Hz, 1H),7.40-7.32 (d, J=9.15, 1H), 7.27-7.18 (t, J=6.04 Hz, 1H), 7.07-6.99 (dd,J=9.15, 2.20 Hz, 1H), 3.89-3.78 (s, 3H), 3.76-3.53 (m, 2H), 3.47-3.30(t, J=6.22 Hz, 2H), 3.07-2.93 (m, 2H), 2.75-2.68 (d, J=10.98 Hz, 2H),2.45-2.24 (m, 4H), 2.07-1.88 (t, J=10.79 Hz, 2H), 1.83-1.59 (m, 7H),1.53-1.35 (m, 2H), 1.18-1.05 (d, J=6.22 Hz, 6H)

LC-MS: 444.30 (MH+)

Synthesis of compound 27—5-methoxy-N-({1-[2-(3-methylcyclohexyl)ethyl]piperidin-4-yl}methyl)-1H-indazole-3-carboxamide

A solution of compound 11b (420 mg, 1.46 mmol) in DMF (45 ml) andtriethylamine (0.61 ml, 4.4 mmol) was stirred at 80° C. for 1 h and thenwas treated with 1-(2-bromoethyl)-3-methylcyclohexane (300 mg, 1.46mmol). The mixture was stirred overnight at the same temperature. Thereaction was then cooled to room temperature and the solvent was removedby evaporation at reduced pressure. The crude5-methoxy-N-({1-[2-(3-methylcyclohexyl)ethyl]piperidin-4-yl}methyl)-1H-indazole-3-carboxamide27 was purified by flash chromatography on silica gel, using a 9/1mixture of CH₃Cl/CH₃OH as eluent. Yield=45 mg (18.0%).

1H NMR (300 MHz, DMSO-d6) δ=13.41 (s, 1H), 8.30-8.20 (t, J=6.11 Hz, 1H),7.58-7.53 (d, J=2.31 Hz, 1H), 7.53-7.47 (d, J=8.59 Hz, 1H), 7.08-7.02(dd, J=8.92, 2.32 Hz, 1H), 3.80 (s, 3H), 3.23-3.13 (t, J=6.28 Hz, 2H),2.90-2.78 (d, J=10.57 Hz, 2H), 2.35-2.20 (m, 2H), 1.97-1.05 (m, 17H),0.90-0.45 (m, 5H)

LC-MS: 413.29 (MH+)

Synthesis of compound28—4-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylicacid

4-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylic acid 28, was prepared,according to the procedure described for compound 25, using methyl4-(chloromethyl)pyridine-2-carboxylate as reagent and CH₃CN as solvent.Yield=335 mg (16%).

¹H NMR (300 MHz, DMSO-d6) δ=13.25 (s, 1H), 8.54 (d, J=4.8 Hz, 1H), 8.27(t, J=6.0 Hz, 1H), 7.93 (s, 1H), 7.56 (d, J=2.2 Hz, 1H), 7.51 (d, J=9.5Hz, 1H), 7.43 (d, J=4.0 Hz, 1H), 7.04 (dd, J=2.2, 9.5 Hz, 1H), 3.80 (s,3H), 3.53 (s, 2H), 3.20 (t, J=6.0 Hz, 2H), 2.78 (d, J=11.0 Hz, 2H), 1.96(t, J=10.6 Hz, 2H), 1.75-1.45 (m, 3H), 1.35-1.16 (m, 2H)

Synthesis of compound 29—Sodium5-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylate29a) Methyl5-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylate

Methyl 5-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylate 29a, was prepared accordingto the procedure described for compound 25 using methyl5-(chloromethyl)pyridine-2-carboxylate as reagent and CH₃CN as solventand used for the subsequent step without further purification.

Then, a solution of crude methyl5-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylate29a (1.2 g, 2.7 mmol) in ethanol (10 ml) was treated with a solution ofNaOH (0.22 g, 5.5 mmol) in water (10 ml) at reflux for 3 h. The mixtureis cooled to room temperature and solvents were evaporated under reducedpressure. The sodium5-{[4-({[(5-methoxy-1H-indazol-3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylate29 was crystallized by a mixture of ethanol/ethyl acetate (1.09 g, 91%).

1H NMR (300 MHz, DMSO-d6) δ=13.86 (br. s., 1H), 8.37 (d, J=1.2 Hz, 1H),8.24 (t, J=6.1 Hz, 1H), 7.91 (d, J=8.1 Hz, 1H), 7.65 (dd, J=2.0, 8.1 Hz,1H), 7.61-7.48 (m, 2H), 7.01 (dd, J=2.5, 8.8 Hz, 1H), 3.79 (s, 3H), 3.47(s, 2H), 3.19 (t, J=6.1 Hz, 2H), 2.77 (d, J=10.9 Hz, 2H), 1.91 (t,J=10.9 Hz, 2H), 1.65 (s, 3H), 1.35-1.07 (m, 2H)

Synthesis of compound30—N-({1-[(5-carbamoyl-1,2,4-oxadiazol-3-yl)methyl]piperidin-4-yl}methyl)-5-methoxy-1H-indazole-3-carboxamide

N-({1-[(5-carbamoyl-1,2,4-oxadiazol-3-yl)methyl]piperidin-4-yl}methyl)-5-methoxy-1H-indazole-3-carboxamide30, was prepared, according to the procedure described for compound 25,using ethyl 3-(chloromethyl)-1,2,4-oxadiazole-5-carboxylate as reagentand CH₃CN as solvent. Yield=80 mg (4%).

¹H NMR (300 MHz, DMSO-d6) δ=13.18 (br. s., 1H), 8.70 (br. s., 1H), 8.32(br. s., 1H), 8.26 (t, J=6.2 Hz, 1H), 7.55 (d, J=1.8 Hz, 1H), 7.50 (dd,J=0.7, 9.1 Hz, 1H), 7.05 (dd, J=1.8, 9.1 Hz, 1H), 3.80 (s, 3H), 3.69 (s,2H), 3.19 (t, J=6.2 Hz, 2H), 2.86 (d, J=11.0 Hz, 2H), 2.19-1.93 (m, 2H),1.82-1.39 (m, 3H), 1.33-1.07 (m, 2H)

Synthesis of compound31—N-({1-[2-(4-nitrophenyl)ethyl]piperidin-4-yl}methyl)-1H-indazole-3-carboxamidehydrochloride

A mixture of 7H,14H-pyrazino[1,2-b:4,5-b′]diindazole-7,14-dione (8.2 g,28.5 mmol), 1-{1-[2-(4-nitrophenyl)ethyl]piperidin-4-yl}methanamine (15g, 57 mmol) in toluene (300 ml) was stirred at room temperature overnight. The solid so obtained was filtered, dissolved with 2N HCl (100ml) and washed with diethylether (3×150 ml). The acid phase was basifiedwith NaOH and extracted with DCM (3×200 ml). The solvent was removed byvacuum and the residue was then poured in THF (30 ml) and treated with1.25 M HCl in MeOH. The crude, solidN-({1-[2-(4-nitrophenyl)ethyl]piperidin-4-yl}methyl)-1H-indazole-3-carboxamidehydrochloride 31 so obtained was filtered and cristallized from EtOH.

¹H NMR (300 MHz, DMSO-d6) δ=13.70 (s, 1H), 10.82 (br. s., 1H), 8.55 (t,J=6.1 Hz, 1H), 8.30-8.10 (m, 3H), 7.67-7.52 (m, 3H), 7.41 (ddd, J=1.2,7.0, 8.4 Hz, 1H), 7.24 (ddd, J=0.8, 7.0, 8.0 Hz, 1H), 3.57 (d, J=11.7Hz, 2H), 3.48-3.15 (m, 6H), 3.04-2.83 (m, 2H), 1.90 (d, J=11.5 Hz, 3H),1.75-1.50 (m, 2H)

The following Table 1A summarizes the chemical name and structure of theabove described compounds 7-31.

TABLE 1A IUPAC name Structure  7 Methyl 2-{[4-({[(5-methoxy-1H-indazol-3- yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-4- carboxylate

 8 2-{[4-({[(5-Methoxy-1H-indazol-3- yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-thiazole-4- carboxylic acid

 9 Methyl 2-{[4-({[(5-methoxy-1H- indazol-3-yl)carbonyl]amino}methyl)piperidin- 1-yl]methyl}-1,3-oxazole-4-carboxylate

10 2-{[4-({[(5-Methoxy-1H-indazol-3- yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}-1,3-oxazole-4- carboxylic acid hydrate

11 Methyl 2-{[4-({[(5-bromo-1H- indazol-3-yl)carbonyl]amino}methyl)piperidin- 1-yl]methyl}-1,3-oxazole-4-carboxylate

12 2-({4-[({[5-(2,3-Difluorophenyl)-1H- indazol-3-yl]carbonyl}amino)methyl]piperidin- 1-yl}methyl)-1,3-oxazole-4-carboxylic acid hydrate

13 Ethyl 4-{[4-({[(5-methoxy-1H- indazol-3-yl)carbonyl]amino}methyl)piperidin- 1-yl]methyl}-1,3-thiazole-2-carboxylate

14 Methyl 2-{[4-({[(5-methoxy-1H- indazol-3-yl)carbonyl]amino}methyl)piperidin- 1-yl]methyl}furan-3-carboxylate

15 Ethyl 5-{[4-({[(5-bromo-1H-indazol- 3-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2- carboxylate

16 5-({4-[({[5-(2-Methoxypyridin-3-yl)- 1H-indazol-3-yl]carbonyl}amino)methyl]piperidin-1-yl}methyl)furan- 2-carboxylic acid hydrate

17 5-({4-[({[5-(6-Methoxypyridin-3-yl)- 1H-indazol-3-yl]carbonyl}amino)methyl]piperidin- 1-yl}methyl)furan-2-carboxylic acidhydrate

18 5-({4-[({[5-(4-Methoxyphenyl)-1H- indazol-3-yl]carbonyl}amino)methyl]piperidin- 1-yl}methyl)furan-2-carboxylic acidhydrate

19 5-({4-[({[5-(2,3-Difluorophenyl)-1H- indazol-3-yl]carbonyl}amino)methyl]piperidin- 1-yl}methyl)furan-2-carboxylic acid

20 5-({4-[({[5-(2-Fluorophenyl)-1H- indazol-3-yl]carbonyl}amino)methyl]piperidin- 1-yl}methyl)furan-2-carboxylic acidhydrate

21 5-({4-[({[5-(4-Methoxypyridin-3-yl)- 1H-indazol-3-yl]carbonyl}amino)methyl]piperidin- 1-yl}methyl)furan-2-carboxylic acidformate

22 5-{[4-({[(5-Bromo-1H-indazol-3- yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylic acid

23 Ethyl 5-{[4-({[(5-methoxy-1H- indazol-3-yl)carbonyl]amino}methyl)piperidin- 1-yl]methyl}furan-2-carboxylate

24 5-{[4-({[(5-Methoxy-1H-indazol-3- yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}furan-2-carboxylic acid

25 N-[(1-{2-[(2R,6S)-2,6- dimethylmorpholin-4-yl]ethyl}piperidin-4-yl)methyl]-5- methoxy-1H-indazole-3- carboxamide

26 N-[(1-{3-[(2R,6S)-2,6- dimethylmorpholin-4-yl]propyl}piperidin-4-yl)methyl]-5- methoxy-1H-indazole-3- carboxamide

27 5-methoxy-N-({1-[2-(3- methylcyclohexyl)ethyl]piperidin-4-yl}methyl)-1H-indazole-3- carboxamide

28 4-{[4-({[(5-methoxy-1H-indazol-3- yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}pyridine-2-carboxylic acid

29 Sodium 5-{[4-({[(5-methoxy-1H- indazol-3-yl)carbonyl]amino}methyl)piperidin- 1-yl]methyl}pyridine-2-carboxylate

30 N-({1-[(5-carbamoyl-1,2,4- oxadiazol-3-yl)methyl]piperidin-4-yl}methyl)-5-methoxy-1H-indazole- 3-carboxamide

31 N-({1-[2-(4- nitrophenyl)ethyl]piperidin-4- yl}methyl)-1H-indazole-3-carboxamide hydrochloride

Pharmacological Properties

The pharmacological properties of the compounds of formula (I) useful inthe present invention were evaluated by the methods described in thefollowing sections.

Test I—Activity on Human GSK-3β (Test In Vitro)

Activity on human GSK-3β was assessed using the following methods(according to Meijer et al., Chem. Biol., 2003-10:1255-1266).

In a first screening assay, compounds were tested in duplicate at aconcentration of 10 μM.

Human recombinant enzyme GSK-3β was incubated for 90 minutes at 22° C.in the presence of compounds or vehicle in a reaction buffer containingATP plus 100 nM unphosphorylated specific substrate peptide(Ulight-CFFKNIVTPRTPPPSQGK-amide). Substrate phosphorylation wasmeasured by LANCE technology (PerkinElmer, Conn., USA).

The results, reported in the following Table 4, are expressed as apercent of inhibition of control specific activity obtained in thepresence of the test compounds (as % inhibition at 10 μm).

In a second assay, the same compounds were assayed at fiveconcentrations ranging from 100 μM to 10 nM with ten-fold dilutions induplicate. Compounds 1 to 7, 9, 11 and 13 to 26 were tested using thesame first assay, compounds 8, 10, 12, and 27 to 31 were tested inanother assay based on the binding and displacement of AlexaFluor® 647labeled, ATP-competitive Kinase inhibitor scaffold using LanthaScreen™TR-FRET technology Eu Kinase assay packet according to manufacturer'sinstruction (Life Technologies, Italy). The results of the two assaysare comparable.

The IC₅₀ values (concentration causing a half maximal inhibition ofcontrol specific activity), reported in table 4, were determined bynon-linear regression analysis of the inhibition curves generated withmean replicate values using Hill equation curve fitting.

TABLE 4 Compound N^(o) % Inhibition [10 μM] IC₅₀ [μM] 1 94 0.35 2 910.56 3 0.31 4 0.64 5 0.35 6 0.40 7 0.13 8 0.17 9 0.21 10 0.31 11 0.36 120.01 13 0.95 14 0.23 15 0.26 16 0.36 17 0.02 18 0.30 19 0.01 20 0.02 210.01 22 0.21 23 0.33 24 0.40 25 1.40 26 2.10 27 0.31 28 0.45 29 0.32 300.31 31 7.20

The results showed that the compounds 1 and 2 according to the presentinvention had good inhibitory activity in this assay: at 10 μM the % ofinhibition is greater that 90% and the IC₅₀ is obtained with less than0.60 μM of each compound.

Most of compounds 3 to 31 according to the present invention showed anIC₅₀ value lower than 1.00 μM. Some of them showed an IC₅₀ value at thelowest concentration of the assay (10 nM). The values of IC₅₀ higherthan 1.00 μM obtained with compounds 25, 26 and 31 are still acceptable.

Test II—Selectivity on GSK-3β (Test In Vitro)

(a) Compound 1 was tested against a panel of 60 kinases in order toassess its selectivity. The assays were chosen taking into considerationthe diversity of assay families.

Tested kinases were representative of following kinase sub-families:

-   -   protein-serine/threonine kinases;    -   protein-tyrosine kinases;    -   other kinases; and    -   atypical kinases.

Human recombinant kinases were incubated in the presence of specificpeptide substrates plus ATP for different times (10, 15, 30, 60 or 90minutes) at 22° C. Phosphorylated substrate was detected by LANCE orHTRF technology (CISBIO, MA, USA).

Compound 1 was tested at 10 μM in duplicate.

The results are expressed as a percent of inhibition of control specificactivity obtained in the presence of the test compound 1 and arereported in the following Table 5.

TABLE 5 % inhibition of control Kinase Kinase values for FamilySub-Family Assay compound 1 Protein- RTK c-Met kinase (h) 0 tyrosine RTKEphA4 kinase (h) 0 kinases RTK EphB2 kinase (h) 0 RTK EphB4 kinase (h) 0RTK FGFR1 kinase (h) 6 RTK FGFR4 kinase (h) 2 RTK IGF1R kinase (h) 5 RTKIRK (h) (InsR) 18 RTK Ret kinase (h) 1 RTK TRKA (h) 10 CTK Abl kinase(h) 0 CTK JAK1 (h) 0 CTK JAK2 (h) 0 CTK Fyn kinase (h) 13 CTK Src kinase(h) 0 Protein CMGC GSK3beta (h) 92 serine/ CMGC DYRK1a (h) 63 threonineCMGC PCTAIRE1 kinase (h) 87 kinases CMGC CDC2/CDK1 (h) (cycB) 31 CMGCCDK2 (h) (cycA) 27 CMGC CDK5/p35 (h) 30 CMGC ERK1 (h) 35 CMGC ERK2 (h)(P42mapk) 33 CMGC p38alpha kinase (h) 0 CMGC p38gamma kinase (h) 2 CMGCp38delta kinase (h) 12 CaMK CHK1 (h) 12 CaMK AMPKalpha 13 CaMK CaMK4 (h)14 CaMK DAPK1 (h) 10 CaMK DCAMKL1 (h) 4 CaMK Pim2 kinase (h) 4 CaMKMAPKAPK2 (h) 0 CaMK MNK2 (h) 1 CaMK PhKgamma 2 (h) 6 CaMK Pim1 kinase(h) 2 CaMK smMLCK (h) (MYLK) 0 AGC GRK3/BARK2 8 (h) (ADRBK2) AGCAkt1/PKBalpha (h) 7 AGC MSK1 (h) 8 AGC PDK1 (h) 8 AGC RSK2 (h) 3 AGC PKA(h) 0 AGC PKCalpha (h) 8 AGC PKCbeta 1 (h) 9 AGC PKCgamma (h) 0 CK1CK1alpha (h) 15 STE PAK1 (h) 4 STE HGK (h) (MAP4K4) 17 STE MEK1/MAP2K1(h) 25 STE TAOK2 (TAO1) (h) 41 TKL DLK1 (h) (MAP3K12) 10 TKL IRAK4 (h) 0Other — IKKalpha (h) 0 kinases — IKKepsilon (h) (IKBKE) 4 — MYT1 kinase(h) 1 — NEK1 (h) 1 — NEK7 (h) 9 — AurA/Aur2 kinase (h) 1 — AurB/Aur1kinase (h) 11 Atypical — mTOR kinase (h) (FRAP1) 0 kinases

Compound 1 was also assayed to determine the IC₅₀ values for threedifferent kinases (PCTAIRE1, DYRK1a, and CDK2) in comparison to Gsk3β.The assay was conducted with the same method described above in test I,second assay. The results are summarized in the following Table 5A.

TABLE 5A IC50 [μM] IC50 [μM] IC50 [μM] IC50 [μM] Compound Gsk3β PCTAIRE1DYRK1a CDK2 1 0.35 1.50 2.90 36.0

Results confirmed that compound 1 had an inhibitory activity on GSK-3βand higher affinity to GSK-3β when compared to the other kinases,showing a good selectivity profile. In fact, the IC₅₀ values of Table 5ashowed a selectivity of compound 1 for Gsk3β better than that forPCTAIRE1, DYRK1a, and CDK2 kinases.

(b) Compounds 7, 12, 21 and 24 were tested against the same panel of 60kinases under the same conditions described above for compound 1.

The results are expressed as a percent of inhibition of control specificactivity obtained in the presence of the test compounds and are reportedin the following Table 6.

TABLE 6 Kinase Sub- Compound Compound Compound Kinase Family FamilyAssay Compound 7 12 21 24 Protein-tyrosine RTK c-Met kinase (h) — — 0 2kinases RTK EphA4 kinase (h) — — 2 0 RTK EphB2 kinase (h) — — 2 0 RTKEphB4 kinase (h) — — 3 0 RTK FGFR1 kinase (h) — — 13 0 RTK FGFR4 kinase(h) — — 0 4 RTK IGF1R kinase (h) — — 0 0 RTK IRK (h) (InsR) 0 0 0 3 RTKRet kinase (h) — — 0 0 RTK TRKA (h) 1 5 4 1 CTK Abl kinase (h) — — 0 0CTK JAK1 (h) — — 10 1 CTK JAK2 (h) — — 2 0 CTK Fyn kinase (h) — — 10 3CTK Src kinase (h) 9 15 0 0 Protein CMGC GSK3beta (h) 96 100 96 94serine/threonine CMGC DYRK1a (h) 88 99 99 59 kinases CMGC PCTAIRE1kinase (h) 2 42 94 1 CMGC CDC2/CDK1 (h) 6 77 99 10 (cycB) CMGC CDK2 (h)(cycA) 48 96 100 36 CMGC CDK5/p35 (h) 21 87 98 16 CMGC ERK1 (h) 31 85 8122 CMGC ERK2 (h) (P42mapk) 33 91 89 35 CMGC p38alpha kinase (h) — — 1 0CMGC p38gamma kinase (h) — — — — CMGC p38delta kinase (h) — — 35 4 CaMKCHK1 (h) — — 1 0 CaMK AMPKalpha — — 70 21 CaMK CaMK4 (h) — — 13 11 CaMKDAPK1 (h) — — 5 15 CaMK DCAMKL1 (h) — — 0 0 CaMK Pim2 kinase (h) — — 5 3CaMK MAPKAPK2 (h) — — 0 0 CaMK MNK2 (h) — — 6 0 CaMK PhKgamma 2 (h) — —0 0 CaMK Pim1 kinase (h) — — 10 2 CaMK smMLCK (h) (MYLK) 26 36 — — AGCGRK3/BARK2 (h) — — 4 0 (ADRBK2) AGC Akt1/PKBalpha (h) — — 0 0 AGC MSK1(h) — — — — AGC PDK1 (h) — — 0 0 AGC RSK2 (h) — — 20 1 AGC PKA (h) — — 30 AGC PKCalpha (h) — — 20 0 AGC PKCbeta 1 (h) — — 0 0 AGC PKCgamma (h) —— 0 0 CK1 CK1alpha (h) — — 2 0 STE PAK1 (h) — — 4 1 STE HGK (h) (MAP4K4)21 98 99 19 STE MEK1/MAP2K1 (h) 27 86 — — STE TAOK2 (TAO1) (h) 16 81 548 TKL DLK1 (h) (MAP3K12) — — 46 0 TKL IRAK4 (h) — — 18 1 Other kinases —IKKalpha (h) — — 5 2 — IKKepsilon (h) — — 21 0 (IKBKE) — MYT1 kinase (h)— — 0 0 — NEK1 (h) 0 30 — — — NEK7 (h) — — 5 3 — AurA/Aur2 kinase (h) —— 30 4 — AurB/Aur1 kinase (h) — — 10 0 Atypical — mTOR kinase (h) — — —— kinases (FRAP1)

Results confirmed that also compounds 7 and 24 had an inhibitoryactivity on GSK-3β and higher affinity to GSK-3β when compared to allother kinases, showing a good selectivity profile, and that compounds 12and 21 had an inhibitory activity on GSK-3β and good affinity to GSK-3βwhen compared to most of other kinases of the same family and to thekinases of different families.

The invention claimed is:
 1. A method of treating a pathological statearising from uncontrolled activation and/or overexpression of GSK-3β,selected from (i) an insulin-resistance disorder, (ii) aneurodegenerative disease, (iii) a mood disorder, (iv) a schizophrenicdisorder, (v) a cancerous disorder, (vi) inflammation, (vii) a substanceabuse disorder, and (viii) an epilepsy, the method comprisingadministering to a human being in need of such treatment an effectiveamount of a 1H-indazole-3-carboxamide according to formula (I) or anaddition salt of said 1H-indazole-3-carboxamide with a pharmaceuticallyacceptable organic or inorganic acid or base:

wherein: each of R_(a) and R_(a)′ is independently selected from ahydrogen atom, a halogen atom, a C₁-C₆ alkyl group, a C₂-C₆ alkenylgroup, a C₂-C₆ alkynyl group, a C₁-C₆ alkoxy group, and an aliphatic oraromatic carbocyclic or heterocyclic ring having from 3 to 12 members,the alkyl, alkenyl, alkynyl, or alkoxy group being optionallysubstituted by at least one substituent selected from a halogen atom, ahydroxyl group, —NH₂, and a C₁-C₃ alkoxy group, and the ring beingoptionally substituted by at least one substituent selected from ahalogen atom, a hydroxyl group, a C₁-C₆ alkyl group, a C₁-C₆ alkoxygroup, —NR₁R₂, —C(O)OH, —C(O)OR₁, and —C(O)NR₁R₂; Y is selected from abond, a C₁-C₆ alkyl group, a C₂-C₆ alkenyl group, and a C₂-C₆ alkynylgroup, the alkyl, alkenyl, or alkynyl group being optionally substitutedby at least one substituent selected from a halogen atom, a hydroxylgroup, —NH₂, and a C₁-C₃ alkoxy group; R_(b) is an aliphatic or aromaticcarbocyclic or heterocyclic ring having from 3 to 12 members,substituted by at least one substituent selected from a halogen atom, ahydroxyl group, a nitro group, a cyano group, —CF₃, a C₁-C₆ alkoxygroup, a benzyloxy group, a C₁-C₄ alkyl group, a C₂-C₄ alkenyl group, aC₂-C₄ alkynyl group, —NHSO₂CH₃, —SO₂NH₂, —Z—C(O)OH, —Z—C(O)OR₁, and—Z—C(O)NR₁R₂; Z is a σ-bond or a (C₁-C₃)alkyl group; and each of R₁ andR₂ is independently selected from a hydrogen atom, a C₁-C₄ alkyl group,a C₂-C₄ alkenyl group, a C₂-C₄ alkynyl group, and a phenyl group.
 2. Themethod of claim 1, comprising treating a neurodegenerative diseaseselected from Parkinson's disease, Alzheimer's disease, Huntington'sdisease, and a spinal neurodegenerative disorder.
 3. The method of claim2, comprising treating a spinal neurodegenerative disorder selected fromamyotrophic lateral sclerosis, multiple sclerosis, spinal muscularatrophy, and neurodegeneration due to spinal cord injury.
 4. The methodof claim 1, comprising treating a mood disorder selected from a bipolardisorder and a depressive disorder.
 5. The method of claim 4, comprisingtreating a bipolar disorder selected from bipolar I, bipolar II,cyclothymia, and a bipolar disorder not otherwise specified (BD-NOS). 6.The method of claim 4, comprising treating a depressive disorderselected from major depressive disorder (MDD), atypical depression (AD),melancholic depression, psychotic major depression (PMD), catatonicdepression, postpartum depression (PPD), seasonal affective disorder(SAD), dysthymia, and a depressive disorder not otherwise specified(DD-NOS).
 7. The method of claim 1, comprising treating an abusedisorder due to psychostimulants.
 8. The method of claim 1, comprisingtreating a schizophrenic disorder selected from paranoid schizophrenia,disorganized schizophrenia, catatonic schizophrenia, simpleschizophrenia, residual schizophrenia, and undifferentiatedschizophrenia.
 9. The method of claim 1, comprising treating a cancerousdisorder selected from prostate cancer, pancreatic cancer, ovariancancer, colon-rectal cancer, and MLL-associated leukaemia.
 10. Themethod of claim 1, comprising treating an insulin-resistance disorderselected from type-2 diabetes, syndrome X, obesity, and polycystic ovarysyndrome. 11.N-{[1-(2,4-dichlorobenzyl)piperidin-4-yl]methyl}-5-methoxy-1H-indazole-3-carboxamide.12.N-({1-[4-(benzyloxy)benzyl]piperidin-4-yl}methyl)-5-methoxy-1H-indazole-3-carboxamide.